Synthesis of 2-[(2-chloro-2′-[18F]fluoroethyl)amino]-2H-1, 3,2-oxazaphosphorinane-2-oxide (18F-fluorocyclophosphamide), a potential tracer for breast tumor prognostic imaging with PET

Article abstract of DOI:10.1002/jlcr.954

A fluorine-18 labeled analog of the widely used chemotherapeutic agent cyclophosphamide was synthesized as a tracer for prognostic imaging with positron emission tomography. 2-[(2-Chloro-2′-[¹⁸F]fluoroethyl) amino]-2H-1,3,2-oxazaphosphorinane-2-oxide (¹⁸F- fluorocyclophosphamide), was prepared by direct halogen exchange reaction from the parent cyclophosphamide. In small-scale syntheses, radiochemical yields of up to 4.9% and specific activities of 960 Ci/mmol were achieved in a total synthesis time of 60-75 min. The [¹⁸F]-labeled cyclophosphamide analog with radioactive purity >99% and chemical purity >96% was suitable for in vivo (microPET imaging) and ex vivo studies of a murine model of human breast tumors. Copyright

Full text of DOI:10.1002/jlcr.954

JOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS
J Label Compd Radiopharm 2005; 48: 635–643.
Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jlcr.954
Research Article
Synthesis of 2-[(2-chloro-20- [18F]fluoroethyl)amino]-
2H-1,3,2-oxazaphosphorinane-2-oxide
(18F-fluorocyclophosphamide), a potential tracer
for breast tumor prognostic imaging with PET
Goran Lacan1,y, Amanda L. Kesner1,y, Anne Gangloff1,z, Lei Zheng2,
Johannes Czernin1, William P. Melega1 and Daniel H. S. Silverman1,*
1 Department of Molecular and Medical Pharmacology, and Ahmanson Biological
Imaging Division, David Geffen School of Medicine, University of California,
Los Angeles, USA
2 Abbott Laboratories, Abbott Park, IL, USA
Summary
A fluorine-18 labeled analog of the widely used chemotherapeutic agent cyclo-
phosphamide was synthesized as a tracer for prognostic imaging with positron
emission
tomography.
2-[(2-Chloro-20-[18F]fluoroethyl)amino]-2H-1,3,2-oxaza-
phosphorinane-2-oxide (18F-fluorocyclophosphamide), was prepared by direct
halogen exchange reaction from the parent cyclophosphamide. In small-scale
syntheses, radiochemical yields of up to 4.9% and specific activities of 960 Ci/mmol
were achieved in a total synthesis time of 60–75 min. The [18F]-labeled cyclo-
phosphamide analog with radioactive purity >99% and chemical purity
>96% was suitable for in vivo (microPET imaging) and ex vivo studies
of a murine model of human breast tumors. Copyright # 2005 John Wiley &
Sons, Ltd.
Key Words: fluorine-18; cyclophosphamide; PET prognostic imaging; breast tumor
therapy
*Correspondence to: D. H. S. Silverman, Nuclear Medicine, CHS AR-144, MC694215, Los Angeles,
California, USA. E-mail: dsilver@ucla.edu
yContributed equally to this work.
zCurrent address: 2538 De Monceaux app 37, Sillery, QC, Canada G1T 2N7.
Contract grant sponsor: California Breast Cancer Research Program Award
Contract grant sponsor: NIH Research Training in Pharmacological Science Training Grant
Contract grant sponsor: University of California Research Mentorship Program
Copyright # 2005 John Wiley & Sons, Ltd.
Received 30 July 2004
Revised 2 September 2004
Accepted 28 March 2005

636
G. LACAN ET AL.
Introduction
Cyclophosphamide (CP) (2-[bis(2-chloroethyl)amino]-2H-1,3,2-oxazaphosphor-
inane 2-oxide) 7 is one of the most widely used agents in cancer chemotherapy.
The report of its synthesis1,2 marked the beginning of rational design and
development of chemotherapeutic drugs.3 It was the first synthetic compound
with a profound anticancer activity in humans. CP itself is a pro-drug, which
undergoes metabolic activation by hepatic cytochrome P450 enzymes to
generate alkylating species.4 Phosphoramidic mustard that is formed in a
series of enzymatic and non-enzymatic transformations is the active species
that crosslinks strands of DNA preventing DNA replication and promoting
cell death. Various by-products of CP biotransformations (e.g. acrolein, and
chloroacetaldehyde) are neurotoxic, urotoxic and cardiotoxic, increasing and
complicating the overall toxicity of chemotherapy.
Cyclophosphamide has been a drug of choice for the treatment of breast
cancer. However, it has been realized that individual patient differences in the
biodistribution, pharmacokinetics, and metabolism of cyclophosphamide and
its analogs (e.g. ifosfamide) can influence the clinical outcome of therapy.5,6
Due to its toxicity, it is particularly worthwhile to assess the likelihood of
benefit from treatment with CP prior to administration of the drug to the
patient. It has been shown that cancer cell lines in vitro can quickly develop
resistance to CP.7ꢀ9 In addition, prior exposure to alkylating agents increases
the likelihood of resistance to CP.7ꢀ11 Both genetic and physiologic alterations
can occur to render the cells resistant to CP.12ꢀ14 An inverse correlation has
been shown between the cellular sensitivity to CP and the cellular content of at
least two aldehyde dehydrogenases, ALDH1A1 and ALDH3A115 that
catalyze the detoxification of CP and its metabolites.16 Glutathione S-
transferase activity, has also been implicated in cellular resistance to CP.8
Information about the biodistribution and kinetics of metabolism of cytotoxic
drugs and CP in particular in individual patients could help to further improve
the selection of appropriate chemotherapeutic regimens.
Positron emission tomography (PET)17ꢀ19 and single photon emission
computed tomography (SPECT)20 have been recently used for the assessment
of the potential response to chemotherapy in patients with breast cancer.
Nuclear medicine imaging modalities allow for the pre-therapy studies of
pharmacokinetic parameters (tissue concentration, times of transit and
retention) in vivo with tracer levels of chemotherapeutics, potentially enabling
a better individualized approach to the administration of pharmacologic doses
of chemotherapy compounds.
Here, we describe the synthesis of a [18F]-labeled analog of cyclopho-
sphamide (8) as a potential tracer for the study of tumor chemosensitivity
in vivo. Since the multistep synthesis of fluorocyclophosphamide21,22 (6) was
not compatible with the rapid time-frame required for a radiosynthesis using a
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SYNTHESIS OF 18F-FLUOROCYCLOPHOSPHAMIDE
637
short-lived isotope such as 18F, we approached labeling through a direct
halogen exchange reaction, starting with a commercially available precursor.23
Results and discussion
In order to identify unequivocally product of the radiosynthetic fluorination,
the desired cold standard (6), a fluorine analogue of cyclophosphamide, was
prepared (Figure 1). The preparation of 6 was based on the modified reported
synthetic procedures.21,22 The multi-step synthesis yielded a racemic 6, an oil
which, unlike cyclophosphamide, did not form crystalline hydrate. Previous
biological testing of this compound has shown its cytotoxicity in HeLa (KB)
cell cultures.22 A 31P NMR study3 of apparent half-lives at pseudo-equilibrium
of 6 and its acyclic metabolite in 1 M lutidine at pH 7.4, 378C has shown that 6
has a longer average apparent half-life compared to cyclophosphamide
(48 ꢁ 1 min vs 38 ꢁ 2 min). At the same time, the relative percentage ratio of
metabolites (cyclic: acyclic) in both cases remains very similar (for 6 it is 84:16
and for cyclophosphamide it is 86:14).
Various synthetic strategies for the halogen exchange reaction were tested in
order to optimize conditions for the [18F]F labeling of 7. For example,
fluorination of 7 with KF/tetrabutylammonium bromide in acetonitrile did
not yield fluoroethyl functionality. However, Kryptofix1222-catalyzed KF
fluorination of anhydrous 724 in acetonitrile25 gave the desired product 6
(Figure 2), whose identity was confirmed by spectroscopic and chromato-
graphic analyses. Chemical shifts and the splitting patterns with the
corresponding proton–fluorine spin–spin coupling values that are character-
1
istic of the fluoroethyl functionality26 were identified in the 19F and H NMR
spectra of the obtained 6. Among the multiple products in this reaction, no
other substances containing fluoroethyl group(s) were identified. Fluorination
with KF/Kryptofix1222 in DMSO at higher temperatures (120–1508C) gave a
mixture of products without formation of 6. Similarly, the attempt to
synthesize the monotosyl congener of cyclophosphamide as a potential
i
ii
FCH2COOC2H5
FCH2CONH(CH2)2OH
FCH2CONH(CH2)2Cl
iii
1
2
3
F(CH2)2NH(CH2)2Cl.HCl
H
N
O
P
iv
4
FCH2CH2
N
FCH2CH2
ClCH2CH2
v
NPOCl2
ClCH2CH2
6
O
5
Figure 1. Synthesis of 2-[(2-chloro-20-fluorodiethyl)amino]-2H-1,3,2-oxazapho-
sphorinane 2-oxide (6): (i) ethanolamine; (ii) SOCl2/CHCl3; (iii) diborane/THF;
(iv) POCl3; (v) 3-aminopropanol, triethylamine/dioxane
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638
G. LACAN ET AL.
H
N
H
N
O
O
i.,ii,iii
CH
2CH2X
P
P
N(CH2CH2Cl)2
N
O
O
CH2CH2Cl
6: X = F
8: X = 18F
7
Figure 2. Direct halogen exchange synthesis of 6 and radiochemical synthesis of
2-[(2-chloro-20-[18F] fluoroethyl)amino]-2H-1,3,2-oxazaphosphorinane-2-oxide
(8): (i) KF or 18Fꢀ/K2CO3/acetonitrile; (ii) Kryptofix1222 1108C/10 min; (iii)
chromatography
substrate for [18F] fluorination, via 7/silver tosylate reaction27ꢀ29 failed to
produce the desired product.
The [18F] analogue of 6 was prepared in a single step by a halogen exchange
reaction with no-carrier-added [18F]Fꢀ/Kryptofix1222 in anhydrous acetoni-
trile with a commercially available cyclophosphamide monohydrate (7)
(Figure 2). Anhydrous cyclophosphamide that was used in the analogous
synthesis of 6 is poorly soluble in cold acetonitrile. An additional evaporation
step upon the addition of cyclophosphamide monohydrate in acetonitrile to
the dried no-carrier-added [18F]Fꢀ/Kryptofix1222 mixture did not improve
radiochemical yield and was omitted in the radiosyntheses. Normal phase
semi-preparative HPLC (Econosil silica column, dichloromethane:methanol,
98:2) was initially tested for the separation and purification of 8, but was
found to require additional filtration and evaporation steps which was
determined to be impractical. A significant difference in the retention times
between the product and the precursor in the reversed phase HPLC, allowed
for their baseline separation. The reversed phase HPLC separation in
combination with a Sep-Pak purification step was used for the work-up of
the reaction mixture. This procedure lends itself to automation in higher
activity syntheses. In small-scale syntheses, starting with 20 mCi 18Fꢀ and a
total synthesis time of 60–75 min, labeling yields (recalculated to the end of
bombardment (EOB)) of up to 4.9% (1.1–4.9%; n ¼ 16) were achieved.
Typically, the activity of injectable product thus obtained (0.35–1.6 mCi) was
sufficient to be used in microPET studies with rodents.
The chromatographic (HPLC, thin-layer chromatography) behavior of the
labeled product 8 was shown to be identical to the unlabeled standard (6)
synthesized in a multi-step procedure and the product obtained in a cold
synthesis with KF/Kryptofix1222. The radiochemical purity of product 8
determined by analytical HPLC was greater than 99% and chemical purity
greater than 96%, with specific activities up to 960 Ci/mmol (54–960 Ci/
mmol).
Copyright # 2005 John Wiley & Sons, Ltd.
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SYNTHESIS OF 18F-FLUOROCYCLOPHOSPHAMIDE
639
Preliminary in vivo experiments (microPET) performed with tumor-bearing
nude mice have shown potential of 8 as a tracer for chemosensitivity study in a
murine model of human breast cancer.
Experimental
Chemicals for the syntheses were purchased from commercial sources (Sigma-
Aldrich, Fisher Scientific) and were used without further purification. All
solvents were of the highest grade commercially available. Tetrahydrofuran
(THF) (Aldrich) was distilled over LiAlH4 under argon. Melting points were
determined with a Thomas–Hoover capillary melting point apparatus and are
uncorrected, as are the reported boiling points. Proton (1H) and 19F spectra
were recorded with a Bruker AM-360 WB spectrometer operating at
360.14 MHz and 338.87 MHz, respectively. Chemical shifts (d, ppm) were
referenced to tetramethylsilane (TMS) for 1H and CFCl3 for 19F as standards.
Coupling constants (J) are reported in Hz. High resolution electron impact (EI
HRMS) mass spectral data were collected on a VG Analytical AutoSpec mass
spectrometer.
Reversed phase analytical high performance liquid chromatography
(HPLC) (Alltech Econosil C18 250 ꢂ 4.6 mm, 5 mm column) and semi-
preparative HPLC (Alltech Econosil C18, 250 ꢂ 10 mm, 5 mm column) were
performed with 0.025 M KH2PO4: acetonitrile (pH 3.0), 75:25 mixture as an
eluent at a flow rate of 1.5 ml/min and 5 ml/min, respectively. The effluent in
semi-preparative HPLC was monitored with an ultraviolet detector at 200 nm
coupled with a gamma-radioactivity PIN diode detector (Carrol and Ramsey
Associates, Berkeley, CA; Model 105-S). For analytical HPLC an UV detector
was coupled with a NaI(Tl)/PMT gamma detector (Bioscan Flowcount,
Model FC-3200). Semi-preparative normal phase HPLC was performed on
Econosil silica 250 ꢂ 10 mm column (Alltech) with either chloroform:methanol
(95:5) or dichloromethane:methanol (98:2). Specific activity of the product 8
was determined by analytical reversed phase HPLC with an UV detector at
200 nm, 0.02 AUF. The linear-response standard curve for the determination
of mass of the carrier product was plotted for the concentrations of 6 between
0.001 and 0.01 mg/ml.
Analytical thin-layer chromatography (TLC) of cyclophosphamide and
fluorinated analogue was carried out on aluminum backed Silica gel 60
(0.25 mm) plates (EM Science) with ethyl acetate:dichloromethane:methanol
(5:3:1). Separated components were visualized by exposure to iodine vapor or
by spraying with 0.2% ninhydrin solution containing 0.15% acetic acid and
heating.30 Flash chromatography was carried out with Silica gel 60, 0.04–
0.063 mm (EM Science).
No-carrier-added [18F]fluoride ion was produced with CTI/Siemens RDS
112 negative ion cyclotron (Knoxville, TN) by the 18O(p,n)18F reaction
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640
G. LACAN ET AL.
with 11 MeV protons in a silver target using [18O]H2O (97% isotopic
enrichment).
N-(2-hydroxyethyl)fluoroacetamide (2)
The title product 2 was obtained as an oil (b.p. 98–1038C; 0.05 mmHg; 89%
yield) (Lit21 b.p. 110–1158C; 0.1–0.2 mmHg). 1H NMR (CDCl3): 3.13 (bs, 1H,
OH); 3.51 (t, 2H, OCH2, JH;H ¼ 5:2); 3.75 (t, 2H, NCH2, JH;H ¼ 5:2); 4.82 (d,
2H, FCH2, JF;H ¼ 47:3); 6.96 (bs, 1H, NH). 19F NMR (CDCl3): ꢀ226.3 (t,
JH;F ¼ 47:0).
N-2-chloroethylfluoroacetamide (3)
The product 3 was synthesized according to the published procedure31 (m.p.
1
65–668C; yield 91%) (Lit31 m.p. 63–658C). H NMR (CDCl3): 3.70 (m, 4H,
CH2CH2); 4.85 (d, 2H, FCH2, JF;H ¼ 47:0); 6.75 (bs, 1H, NH). 19F NMR
(CDCl3): ꢀ226.5 (t, JH;F ¼ 47:0).
N-(2-chloroethyl)-N-(2-fluoroethyl)amine hydrochloride (4)
Diborane reduction of amide 321 yielded after recrystallization from
anhydrous acetone a crystalline 4 (m.p. 193–1948C; yield 78%) (Lit21 m.p.
1
193–1958C). H NMR (D2O): 3.49 (t, 2H, NCH2-(CH2F) JH;H ¼ 4:6); 3.57
(m, 2H, CH2Cl); 3.94 (t, 2H, NCH2-(CH2Cl), JH;H ¼ 5:6); 4.8 (dt, 2H,
CH2F, JH;H ¼ 4:6, JF;H ¼ 46:7). 19F NMR (CDCl3): ꢀ223.5 (2JH;F ¼ 46:7,
3JH;F ¼ 29:3).
N-(2-chloroethyl-N-(2-fluoroethyl)phosphoramidic dichloride (5)
The product 5 was obtained22 as a colorless oil (b.p. 97–1038C; 0.5 mmHg;
yield 47%) (Lit.22 b.p. 97–998C at 0.4–0.5 mmHg). 1H NMR (CDCl3): 3.66 (m,
6H, 3CH2); 4.66 (dt, 2H, FCH2, JF;H ¼ 47:3:0 Hz, JH;H ¼ 4:6 Hz). 19F NMR
3
(CDCl3): ꢀ222.6 (2JH;F ¼ 47:3 Hz, JH;F ¼ 29:3 Hz).
2-[(2-Chloro-20-fluorodiethyl)amino]-2H-1,3,2-oxazaphosphorinane-2-oxide (6)
The condensation of 5 and 3-aminopropanol afforded 6 as a colorless viscous
oil22 (yield 84%). A portion of the product (740 mg) was dissolved in
dichloromethane and flash-chromatographed on a silica gel column with
dichloromethane:methanol, 18:1. Fractions were examined with TLC, and
ninhydrin positive fractions with Rfꢃ0.3 were pooled and evaporated under
reduced pressure. Colorless oil was obtained (612 mg, 83%). 1H NMR
5
(CDCl3): 1.86 (m, 2H, CH2); 2.63 (bs, 1H, NH); 3.44 (m, 6H, N-CH2-(F);
4
N-CH2-(Cl), CH2); 3.65 (t, 2H, CH2Cl, JH;H ¼ 7:0); 4.36 (m, 2H,6CH2); 4.55
(dt, 2H, FCH2, JF;H ¼ 47:3:0, JH;H ¼ 4:9). 19F NMR (CDCl3): ꢀ222.6
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SYNTHESIS OF 18F-FLUOROCYCLOPHOSPHAMIDE
641
(2JH;F ¼ 47:3, 3JH;F ¼ 29:3); EI HRMS M++H calculated for C7H16N2O2FPCl:
245.0622, found: 245.0618.
2-[(2-Chloro-20-fluorodiethyl)amino]-2H-1,3,2-oxazaphosphorinane-2-oxide
(6) by direct fluorination of (7)
Anhydrous cyclophosphamide24 7 (20 mg, 0.08 mmol) was heated at reflux in
anhydrous acetonitrile (2.5 ml) (oil bath temperature 95–1008C) in the
presence of Kryptofix1222 (78 mg, 0.21 mmol) and anhydrous potassium
fluoride (11 mg, 0.19 mmol) for up to 3 h. Thin layer chromatography of the
reaction mixture showed the appearance of multiple products. 19F NMR
(CDCl3) of the reaction mixture indicated the presence of a signal at ꢀ221 ppm
that is characteristic of the fluoro-alkyl functionality. The reaction mixture
was evaporated under reduced pressure and dissolved in dichloromethane.
Chromatography on a silica gel column with dichloromethane:methanol (18:1)
1
afforded the product 6 (2 mg; 10.7%) whose 19F and H NMR spectra
were identical with the standard synthesized by the above described methods.
The product obtained by the KF/Kryptofix1222 fluorination was
co-chromatographed on analytical reversed phase HPLC with the standard
6 (Rt 6 ¼ 4:5 min; Rt 7 ¼ 7:5 min).
2-[(2-Chloro-20-[18F]fluorodiethyl)amino]-2H-1,3,2-oxazaphosphorinane-2-oxide
(18F-fluorocyclophosphamide)(8)
[18F]-fluorocyclophosphamide (8) was prepared by a single-step halogen
exchange as shown in Figure 2. Typically, [18F]Fꢀ recovered in water (300 ml)
was added to a 0.1 M solution of potassium carbonate (30 ml) and
Kryptofix1222 (2.25 mg, 6 mmol) in 50 ml acetonitrile) and evaporated to
dryness at 1108C in a stream of argon. Acetonitrile was added (3 ꢂ 300 ml) and
evaporated to dryness. The solution of cyclophosphamide monohydrate (6 mg,
21 mmol)) in acetonitrile (600 ml) was then added and the reaction mixture
heated at 1108C in a closed vial for 10 min. The reaction mixture was, after
brief (3 min) cooling to room temperature in an ice bath, directly injected onto
a semi-preparative reversed phase HPLC column eluted with a flow rate of
5 ml/min. The radioactive fraction at 7.7–8.7 min was collected (Rt 8=8.2 min;
Rt cyclophosphamide (7)=12.8 min). pH of the collected eluate was adjusted
to 7.8 with 0.025 M NaH2PO4 and diluted tenfold with water (total volume
ꢃ50 ml). The diluted eluate was passed through a preconditioned C18 Sep-
Pak column (Waters, C18 Sep-Pak, 300 mg), activated with methanol (5 ml)
and water (10 ml). The column was then washed with water (25 ml). The
radioactive product 8 was eluted with acetonitrile (1 ml). The acetonitrile was
evaporated under reduced pressure, the residue was dissolved in ethanol (1 ml)
and then evaporated to dryness (T5358C). The injectable radioactive tracer
was obtained after reconstitution in 0.9% saline and filter-sterilization
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G. LACAN ET AL.
(0.22 mm filter, Millex1GP, PES membrane, Millipore). Radiochemical and
chemical purities were determined by analytical HPLC and were found to be
>99% and >96%, respectively.
Conclusion
We have synthesized an analog of cyclophosphamide that contains a single
[18F]-fluoroethyl group (‘fluorocyclophosphamide’) by direct halogen ex-
change reaction from a commercially available precursor. The resulting
radioactive tracer, of high radiochemical and chemical purity, is suitable for
in vivo (microPET) studies of pharmacokinetic parameters relevant to
individualized pre-therapeutic assessment of therapy regimens.
Acknowledgements
The authors gratefully acknowledge Prof. J.R. Barrio, Prof. N. Satyamurthy
and V. Kepe PhD of UCLA Department of Molecular and Medical
Pharmacology for advice, suggestions and helpful discussions and UCLA
Biomedical Cyclotron staff for the production of the 18F isotope. This work
was supported by the California Breast Cancer Research Program award.
A.L.K. was supported by the NIH Research Training in Pharmacological
Sciences Training Grant and University of California Research Mentorship
Program.
References
1. Arnold H, Bourseaux F. Agnew Chem 1958; 70: 539–544.
2. Arnold H, Bourseaux F, Brock N. Nature 1958; 181: 931.
3. Ludeman SM. Curr Pharm Des 1999; 5: 627–643.
4. Malet-Martino M, Gilard V, Martino R. Curr Pharm Des 1999; 5: 561–586.
5. Sreerama L, Sladek NE. Cancer Chemother Pharmacol 2001; 47: 255–262.
6. Sladek NE, Kollander R, Sreerama L, Kiang DT. Cancer Chemother Pharmacol
2002; 49: 309–321.
7. Wu L, Tannock IF. Cancer Res 2003; 63: 2134–2138.
8. Gamcsik MP, Dolan ME, Andersson BS, Murray D. Curr Pharm Des 1999; 5:
587–605.
9. Teicher BA, Herman TS, Holden SA, Wang YY, Pfeffer MR, Crawford JW, Frei
E 3rd. Science 1990; 247: 1457–1461.
10. Graham CH, Kobayashi H, Stankiewicz KS, Man S, Kapitain SJ, Kerbel RS. J
Natl Cancer Inst 1994; 86: 975–982.
11. Teicher BA, Kakeji Y, Ara G, Herbst RS, Northey D. In vivo 1997; 11: 453–461.
12. Kobayashi H, Man S, Graham C, Kapitain SJ, Teicher BA, Kerbel RS. Proc Natl
Acad Sci USA 1993; 90: 3294–3298.
13. Begg AC, Smith KA. Br J Cancer 1984; 49: 49–55.
14. Houghton PJ, Tew KD, Taylor DM. Cancer Treat Rep 1976; 60: 459–464.
15. Sladek NE. Curr Pharm Des 1999; 5: 607–625.
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 635–643

SYNTHESIS OF 18F-FLUOROCYCLOPHOSPHAMIDE
643
16. Moreb J, Zucali JR, Zhang Y, Colvin OM, Gross MA. Cancer Res 1992; 52:
1770–1774.
17. Varagnolo L, Stokkel MPM, Mazzi U, Pauwels EKJ. Nucl Med Biol 2000; 27:
103–112.
18. Silverman DH, Hoh CK, Seltzer MA, Schiepers C, Cuan GS, Gambhir SS, Zheng
L, Czernin J, Phelps ME. Semin Radiat Oncol 1998; 8: 183–196.
19. Pio BS, Park CK, Satyamurthy N, Czernin J, Phelps ME, Silverman DH. J Nucl
Med 2003; 44: 76P–77P.
20. Takeuchi N, Fukumoto M, Nishioka A, Akagi N, Murata Y, Takeuchi S, Ogawa
Y, Yoshida S. Int J Oncol 2002; 20: 53–58.
21. Papanastassiou Z, Bruni J. J Org Chem 1964; 29: 2870–2872.
22. Papanastassiou Z, Bruni J, Potts Fernandes F, Levins P. J Med Chem 1966; 9:
357–359.
23. Kesner AL, Lacan G, Gangloff A, Zheng AL, Czernin J , Melega WP, Silverman
DH. J Nucl Med 2003; 44 (5 Suppl): 297P.
24. Reid JM, Stobaugh JF, Sternson LA. Anal Chem 1989; 61: 441–446.
25. Liotta CL, Harris PH. J Am Chem Soc 1974; 96: 2250–2252.
26. Weigert FJ. J Org Chem 1980; 45: 3476–3483.
27. Takahashi T, Mizuno T, Ido T, Iwata T, Watanabe K. Appl Radiat Isot 2003; 58:
557–566.
28. Yoh S.-D, Tsuno Y, Fujio M, Sawada M, Yukawa Y. J Chem Soc Perkin Trans II
1989; 7–13.
29. Kornblum N, Jones WJ, Anderson GJ. J Am Chem Soc 1959; 81: 4113–4114.
30. Tonelli D, Gattavecchia E, Breccia A. J Chromatogr 1983; 275: 223–228.
31. Buckle F, Heap R, Saunders B. J Chem Soc 1949; 912–916.
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 635–643