Design of bisphosphonate-terminated dendrimers

Article abstract of DOI:10.1002/ejoc.200901291

Several new methods for the synthesis of phosphorus dendrimers capped with biologically active azabisphosphonates are reported. Monomers bearing either azido or acetylenic groups were designed as building blocks either for attachment onto the surfaces of

Full text of DOI:10.1002/ejoc.200901291

FULL PAPER
DOI: 10.1002/ejoc.200901291
Design of Bisphosphonate-Terminated Dendrimers
Emma Cavero,[a] Maria Zablocka,[b] Anne-Marie Caminade,*[a] and Jean Pierre Majoral*[a]
Keywords: Dendrimers / Click chemistry / Multistep synthesis / Phosphazenes / Phosphorus
Several new methods for the synthesis of phosphorus dendri-
mers capped with biologically active azabisphosphonates are
reported. Monomers bearing either azido or acetylenic
groups were designed as building blocks either for attach-
ment onto the surfaces of classical phosphorus dendrimers or
for use in the construction of original dendrimers without the
need for use of sensitive dichloro(monomethylhydrazino)-
thiophosphane.
Introduction
Structural features of dendrimers (their nanometer sizes,
perfectly defined structures, topologies, multivalent charac-
ters, molecular weights), together with their high density of
chemical reactive functions on their outer shells explain
their successful use in biology and medicine.[1–8] Among
them, phosphorus-containing dendrimers appear to be ver-
satile macromolecules offering a large palette of properties
and applications. Those bearing phosphonate end groups
are of particular interest. Indeed these dendrimers – such
as the first-generation dendrimer 1 – were found to act as
new nano-biotools promoting anti-inflammatory and im-
munosuppresive activation of human monocytes and so
show promise as drugs for the treatment of uncontrolled
inflammatory process in acute or chronic diseases such as
psoriasis, rheumatoid arthritis or autoimmune disor-
ders.[9,10] Moreover, phosphonates grafted onto the surfaces
of phosphorus dendrimers display the unexpected property
of dramatically and selectively promoting the multiplication
of human natural killer (NK) cells, which are a key part of
innate immunity.[11,12] The exceptional bioactivity behav-
iour of these dendrimers prompted us not only to diversify
with regard to ways of attaching phosphonates onto the
surfaces of known phosphorus dendrimers but also to se-
arch for new ways to synthesise phosphorus dendrimers
possessing terminal phosphonate groups in order to opti-
mize their properties and to find the best molecule for the
applications discussed above.
Here we report: i) the synthesis of monomers as new
building blocks for the construction of dendrimers, ii) the
attachment of some of these building blocks onto new or
“classical” phosphorus dendrimers developed in our team
through “click reactions”, and iii) a strategy of elaboration
of original dendrimers which allows the user to avoid the
use of toxic methylhydrazine and the preparation of dichlo-
ro(methylhydrazino)thiophosphane, which takes place un-
der drastic conditions (–60 °C). This paper focuses on the
preparation of analogues of the dendrimer 1, which ap-
peared to be the most efficient dendrimer for the biomedical
applications discussed above. However, the strategies used
can be extended to dendrimers of higher generations, as is
illustrated below.
Results and Discussion
Various building blocks were prepared for the above
goals. Firstly, acetylenic moieties or azido groups were in-
troduced at the level of the core through the formation of
the hexafunctionalized compounds 3–5 (Scheme 1). Sec-
ondly, monomers bearing either an acetylenic group, as in
compounds 6, or an azido unit, as in compounds 8, were
prepared for grafting onto the surfaces of dendrimers
(Scheme 2).
The high-yield syntheses of the cores 3 and 4 involve nu-
cleophilic substitution between hexachlorocyclotriphos-
phazene – (N3P3)Cl6 – and phenols substituted in their para
positions either with O(CH2)2Cl or with CH2OH groups;
the terminal chlorine or OH groups were then converted
into azido groups. Notably, the use of a phenol bearing a
halogen allows the one-step transformation into the corre-
sponding azide 3 (2a Ǟ 3), whereas use of the 4-hy-
[a] Laboratoire de Chimie de Coordination (LCC) CNRS, and
Université de Toulouse, UPS, INP, LCC,
205 route de Narbonne, 31077 Toulouse, France
Fax: +33-5-61553003
E-mail: caminade@lcc-toulouse.fr
majoral@lcc-toulouse.fr
[b] Centre of Molecular and Macromolecular Studies, Polish Acad-
emy of Sciences,
Sienkiewicza 112, 90363 Łodz, Poland
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propargylamine or 3-aminopropan-1-ol with the phos-
phonate (MeO)2P(O)H and formaldehyde afford the mono-
mers 6 or 7 in high yields. Transformation of the alcohol
derivative 7 into the corresponding azide 8 occurs under
mild conditions (Scheme 2).
The only previously reported way to introduce an amino-
bisphosphonate moiety onto the surface of a first-genera-
tion phosphorus dendrimer was through a reaction between
a terminal P(S)Cl2 group and a phenol such as 9.[12] A new
alternative involves the use of “click chemistry” with den-
drimers decorated with azido groups and monomers bear-
ing acetylenic groups.[13–15]
To test this reaction, in a first approach we treated the
hexaazido core 3 with the acetylenic aminobisphosphonate
6 (Scheme 3) in the presence of CuI and diisopropylethyl-
amine (DIPEA) as base in THF at 40 °C. Under these con-
ditions the expected dendrimer 10-[G0] (generation zero),
incorporating six azabisphosphonate end groups, was
formed. The reaction can be easily monitored by NMR and
IR spectroscopy: the disappearance of the N3 band at
2106 cm–1 is observed by IR spectroscopy, whereas 1H
NMR shows the shift of the N methylene signal from
4.33 ppm for 3 to 5.56 ppm in 10-[G0], as well as the appear-
ance of characteristic signals due to the CH groups of the
triazole units at δ = 7.83 ppm. Moreover, a shielding effect
is detected by 31P NMR for the terminal bisphosphonate (δ
=25.9 ppm for 6, 26.5 ppm for 10-[G0]).
Scheme 1. Synthesis of the azido and acetylenic cores 2a–c and
3–5.
The same sequence of reactions for the introduction of
azido groups onto the surfaces was applied to dendrimers
of the first and second generations bearing P(S)Cl2 end
groups (i.e., dendrimers 11-[G1] and 11-[G2]), leading suc-
cessively to the dendrimers 12-[G1] and 12-[G2] and then 13-
[G1] and 13-[G2]; the latter were successfully treated with
the monomer 6 to give the dendrimers 14-[G1] and 14-[G2],
incorporating 12 and 24 azabisphosphonate linkages,
respectively, thus offering diversity in relation to 1 with re-
Scheme 2. Synthesis of phosphonate building blocks 6–8.
droxybenzyl alcohol requires two steps to prepare the azido gard to the nature of the spacers between the dendrimer
species 4 (2b Ǟ 2c Ǟ 4). Grafting of acetylenic units to backbone itself and the terminal bisphosphonate
(N3P3)Cl6 also proceeds in a one-step process giving rise to (Scheme 3). As in the case of the synthesis of 10-[G0], reac-
the core 5 (Scheme 1). Kabachnick–Field reactions of either tions can be monitored by NMR spectroscopy. For 14-[G1],
Scheme 3. Synthesis of bisphosphonate-ended dendrimers from the azido dendrimers 3, 13-[G1] and 13-[G2] and the acetylenic azabisphos-
phonate 6.
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Design of Bisphosphonate-Terminated Dendrimers
for example, 1H NMR shows the disappearance of the monitored by 31P NMR, which shows a signal at δ =
methylene signal (CH2N3) at δ = 3.54 ppm (t, JH,H
=
26.8 ppm in addition to the other unchanged characteristic
4.8 Hz) and the appearance of a triplet (JH,H = 4.8 Hz) at signals of the phosphorus dendritic backbone. H and 13C
δ = 4.71 ppm, whereas the characteristic signal of the CH NMR spectroscopy as well as IR spectroscopy corroborate
group of the triazole appears as a singlet at δ = 7.82 ppm. the full achievement of the sequence of formation and isola-
Moreover, the methylene group connecting the triazole ring tion of all the acetylenic- or bisphosphonate-terminated
and the azabisphosphonate appears as a singlet at δ = dendrimers.
1
4.15 ppm. 31P NMR spectroscopic data corroborate the
Except for the formation of the dendrimers of genera-
presence of terminal phosphonates whereas 13C NMR and tion zero – 10-[G0] and 15-[G0] – the two approaches re-
IR supply definite confirmation of the formation of the de- ported above do not allow the use of monomethylhydrazine
sired dendrimers.
for the construction of higher generations to be avoided,
A second general alternative for incorporation of azabis- the CH=N–N(CH3)P(S) fragment being present on each
phosphonate units was investigated. It involves the prelimi- generation. Therefore a third general alternative was elabo-
nary grafting of acetylenic functions (a type of functional rated. This involves a multistep synthesis as shown in
group rarely positioned at the surfaces of phosphorus den- Scheme 5. The three first steps necessitated the design of
drimers[16,17]) onto the surfaces of dendrimers, followed by the AB5 monomer 18, obtained through the selective penta-
treatment of the resulting dendrimers with the multifunc- substitution of the hexachlorocyclotriphosphazene P3N3Cl6
tionalized monomer 8 (Scheme 4). As in the first strategy with 4-hydroxybenzyl alcohol and nucleophilic substitution
reported above, such an experiment was conducted with of the remaining chlorine atom with 4-(prop-2-yn-1-yloxy)-
two monomers: the hexaacetylenic monomer 5 and the az- phenol to afford the further AB5 monomer 19, which can
ide 8. This reaction also takes place under mild conditions, be treated with the hexa-azido monomer 3 to give the den-
allowing us to isolate the new generation zero dendrimer, drimer 20-[G1]. In contrast with what is observed with
15-[G0], bearing spacers different from those in its congener phosphorus dendrimers incorporating P(S)N(CH3)N=CH–
10-[G0] on each side of the triazole moieties. This transfor- linkages, no fragmentation is detectable by mass spectrome-
mation can be monitored by NMR spectroscopy: 1H NMR try for such a dendrimer[18] in which the core and the
confirms the total disappearance of the signals due to the branching points are cyclotriphosphazene units. Introduc-
methylene acetylenic linkage at 4.68 ppm (d, JH,H = 2.4 Hz, tion of azido groups onto the outer shell then proceeds as
CH2) and 2.55 ppm (t, JH,H = 2.4 Hz, CH) with the appear- above, with the formation of 21-[G1] (30 terminal chlorines)
ance of singlets at 5.13 ppm (CH2) and at 7.48 ppm (CH, and then of 22-[G1], a dendrimer bearing 30 azido groups.
triazole), as well as all signals attributable both to the Aminobisphosphonates can be grafted onto 22-[G1]
(CH2)3 linkage and to the azabisphosphonate fragments. through [1+3] cycloaddition with the acetylenic monomer
Extension of this method to dendrimers of higher genera- 6, leading to 23-[G1].
tion first necessitates the synthesis of dendrimers bearing
1H NMR spectroscopy nicely illustrates the dissymmetry
acetylenic units on their outer shells. These dendrimers – of this dendrimer through the AB5 system present in 19.
16-[G1] and 16-[G2], prepared through nucleophilic substi- Whereas the CH proton of the triazole ring incorporated in
tution of the terminal P(S)Cl2 groups of dendrimers 11-[G1] the first generation gives a singlet at 8 ppm, those of the
and 11-[G2] with 4-(prop-2-yn-1-yloxy)phenol in the pres- triazole units of the second generation appear as two sing-
ence of caesium carbonate – were then allowed to react with lets in a 2/3 ratio at δ = 7.71 and 7.78 ppm. This is due to
the azido monomer 8 to give the new dendrimers 17-[G1] the presence of the cyclotriphosphazene ring used as
and 17-[G2] (Scheme 4). Completion of the reactions was branching point, allowing a dissymmetric anchoring of the
Scheme 4. Synthesis of bisphosphonate-ended dendrimers from the acetylenic dendrimers 5, 15-[G1] and 15-[G2] and the azido azabisphos-
phonate 8.
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FULL PAPER
Scheme 5. Synthesis of phosphorus dendrimers incorporating triazole units in each generation.
five remaining branches as we have already reported.[17]
Two singlets at δ = 5.54 and 5.51 ppm are observed for the
methylene groups of the C6H4CH2N linkages of the outer
shell in the same 2/3 ratio, as well as for the methylene
groups of the C–CH2–N fragments (4.14 and 4.10 ppm),
whereas the signals for the NCH2P groups appear as dou-
blets at 3.13 and 3.16 ppm (2JH,P = 10 Hz).
spectra were measured at 25 °C in the indicated deuterated solvents.
Proton and carbon chemical shifts (δ) are reported in ppm and
coupling constants (J) are reported in Hertz (Hz). The signals in
the spectra are described as s (singlet), d (doublet), t (triplet), m
(multiplet), br. (broad resonances), and Ctz (triazole signals). Refer-
ences for NMR chemical shifts are H3PO4 (85%) for 31P NMR,
1
and SiMe4 for H and 13C NMR spectroscopy. Signal attribution
was carried out with the aid of two-dimensional experiments when
necessary (COSY, HSQC). The numbering used for NMR signals
assignment in shown in Figure 1. Fourier transformed infrared
Several key features can be emphasized with regard to
23-[G1]: i) it does not contains methylhydrazino units, in
marked contrast with the great majority of the known phos- (FTIR) spectra were obtained with a Perkin–Elmer Spectrum 100
FT-IR spectrometer on neat samples (ATR FT-IR). Mass spec-
trometry was carried out with a Thermo Fisher DS QII (DCI/
NH3), a Neromag R10–10 (FAB) and an Applied Biosystems Voy-
ager System 4243 (MALDI). Either protonated molecular ions [M
+ H]+ or sodium adducts [M + Na]+ were used for empirical for-
mula confirmation.
phorus dendrimers, ii) each generation incorporates triazole
rings, iii) on moving from the first generation to the second
the number of triazole rings is multiplied by five (six for the
first generation, thirty for the second), and iv) it illustrates
the versatility of azide reactivity towards phosphorus com-
pounds, leading either to Staudinger-type reactions[19–22]
with removal of N2 or to [1+3] cycloaddition with retention
of the N3 framework.
Chemicals were purchased from Aldrich, Acros, Fluka and Strem
and were used without further purification, except for P3N3Cl6,
which was recrystallized from hexanes. Organic solvents were dried
and distilled by routine procedures. Purifications by flash column
chromatography were performed with silica gel 60 (43–63 µm).
TLCs were performed on silica gel 60 F254 plates and detection
was carried out either under UV light or with molybdate solution
followed by heating.
Conclusions
New strategies for the synthesis of phosphorus dendri-
mers and new methods for their incorporation on their sur-
faces of biologically active bisphosphonates are reported,
thus expanding the potential to diversify not only their bio-
medical applications but also their involvement in the de-
sign of hybrid organic-inorganic materials.[23] Uses of all
the dendrimers reported here for these applications are un-
der active investigation.
Compound 2a: Cs2CO3 (38 mmol) was added to an acetone (dry,
60 mL) solution of 4-(2-chloroethoxy)phenol (19 mmol) and
P3N3Cl6 (2.88 mmol), and the mixture was stirred at 40 °C until
the reaction was complete (monitored by 31P NMR). Inorganic
salts were filtered through a Celite® pad and the residual solvent
was removed. The crude product was purified by silica gel flash
chromatography (eluent: pentane/CH2Cl2 3:7). White powder; yield
1
75% (2.152 g, 2.16 mmol). H NMR (300 MHz, CDCl3): δ = 6.85
Experimental Section
(d, J = 9.0 Hz, 12 H, H2), 6.72 (d, J = 9.0 Hz, 12 H, H3), 4.20 (t,
J = 5.7 Hz, 12 H, OCH2), 3.82 (t, J = 5.7 Hz, 12 H, CH2Cl) ppm.
13C{1H} NMR (100 MHz, CDCl3): δ = 155.2 (C4), 144.7 (C1),
Methods and Materials: NMR spectra were recorded with Bruker
ARX 250, DPX 300, AV 300, AV 400 or AV 500 spectrometers. All
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Design of Bisphosphonate-Terminated Dendrimers
Figure 1. Numbering used for assignment of NMR signals.
122.0 (C2), 115.4 (C3), 68.6 (CH2O), 42.1 (CH2Cl) ppm. 31P{1H} NMR (300 MHz, CDCl3): δ = 7.24 (d, J = 8.4 Hz, 12 H, H3), 6.93
NMR (101 MHz, CDCl3): δ = 9.9 ppm. MS (FAB): m/z = 1163
[M]+. C48H48Cl6N3O12P3 (1164.6): calcd. C 49.51, H 4.15, N 3.61;
found C 49.38, H 4.14, N 3.54.
(d, J = 8.4 Hz, 12 H, H2), 4.58 (s, 12 H, CH2Cl) ppm. 13C{1H}
NMR (75 MHz, CDCl3): δ = 150.2 (m, C1), 134.3 (C4), 129.9 (C3),
121.2 (C2), 45.6 (CH2Cl) ppm. 31P{1H} NMR (121.5 MHz,
CDCl3): δ = 8.7 ppm. C42H36Cl6N3O6P3 (984.4): calcd. C 51.25, H
3.69, N 4.27; found C 51.10, H 3.62, N 4.19.
Compound 3: NaN3 (0.6 mmol) was slowly added to a DMF (dry,
5 mL) solution of compound 2a (0.5 mmol), and the reaction mix-
ture was stirred and heated at 70 °C overnight. The reaction mix-
ture was then allowed to cool to room temp., distilled water (5 mL)
was added, and a white precipitate appeared. The aqueous phase
was decanted and the precipitate was dissolved in CH2Cl2. The
organic phase was then washed with brine, dried with MgSO4 and
concentrated to give the azide derivative 3, which was further puri-
fied by silica gel flash chromatography (eluent: pentane/CH2Cl2
7:3). Waxy, white solid; yield 85% (0.512 g, 0.425 mmol). 1H NMR
Compound 4: NaN3 (0.6 mmol) was slowly added to a DMF (dry,
5 mL) solution of the chloride derivative 2c (0.5 mmol), and the
reaction mixture was stirred at room temp. overnight. Distilled
water (5 mL) was then added to the reaction mixture and a white
precipitate appeared. The aqueous phase was decanted and the pre-
cipitate was dissolved in CH2Cl2. The organic phase was washed
with brine, dried with MgSO4 and concentrated to give the azide
derivative 4, which was further purified by silica gel flash
(400 MHz, CDCl3): δ = 6.86 (d, J = 8.8 Hz, 12 H, H2), 6.74 (d, J chromatography (eluent: pentane/CH2Cl2 8:2). Waxy, white solid;
= 8.8 Hz, 12 H, H3), 4.12 (t, J = 4.8 Hz, 12 H, OCH2), 3.61 (t, J
= 4.8 Hz, 12 H, CH2N3) ppm. 13C{1H} NMR (100 MHz, CDCl3):
δ = 155.3 (C4), 144.7 (m, C1), 122.0 (C2), 115.2 (C3), 67.5 (OCH2),
yield 89% (0.456 g, 0.445 mmol). 1H NMR (300 MHz, CDCl3): δ
= 7.17 (d, J = 8.7 Hz, 12 H, H3), 6.99 (d, J = 8.7 Hz, 12 H, H2),
4.33 (s, 12 H, CH2N3) ppm. 13C{1H} NMR (100 MHz, CDCl3): δ
50.2 (CH2N3) ppm. 31P{1H} NMR (162 MHz, CDCl3): δ
=
=
150.4 (m, C1), 132.3 (C4), 129.4 (C3), 121.3 (C2), 54.1
9.85 ppm. IR (neat): ν = 2106 (N ) cm–1. MS (FAB): m/z = 1204
(CH2N3) ppm. 31P{1H} NMR (121.5 MHz, CDCl3): δ = 8.4 ppm.
˜
3
[M + 1]+. C48H48N21O12P3 (1204.0): calcd. C 47.89, H 4.02, N
IR (neat): ν = 2090 (N ) cm–1. MS (FAB): m/z = 1023 [M]+.
˜
3
24.43; found C 47.86, H 4.00, N 24.39.
C42H36N21O6P3 (1023.8): calcd. C 49.27, H 3.54, N 28.73; found C
49.18, H 3.51, N 28.67.
Compound 2b: Cs2CO3 (38 mmol) was added to a THF (dry,
60 mL) solution of 4-hydroxybenzyl alcohol (19 mmol) and
P3N3Cl6 (2.88 mmol), and the mixture was stirred at 40 °C until
reaction was complete (monitored by 31P NMR). Inorganic salts
were filtered through a Celite® pad and the residual solvent was
removed. The crude product was purified by silica gel flash
chromatography (eluent: CHCl3/MeOH 9:1). White powder; yield
83% (2.088 g, 2.39 mmol). 1H NMR (400 MHz, [D8]THF): δ =
7.18 (d, J = 8.4 Hz, 12 H, H3), 6.90 (d, J = 8.4 Hz, 12 H, H2), 4.57
(d, J = 6.0 Hz, 12 H, CH2OH), 4.38 (t, J = 6.0 Hz, 6 H,
CH2OH) ppm. 13C{1H} NMR (100 MHz, [D8]THF): δ = 150.1
(C1), 136.2 (C4), 127.2 (C3), 120.4 (C2), 63.3 (CH2OH) ppm.
Compound 5: Cs2CO3 (38 mmol) was added to a THF (dry, 60 mL)
solution of 4-(prop-2-yn-1-yloxy)phenol[24] (19 mmol) and P3N3Cl6
(2.88 mmol), and the mixture was stirred at 40 °C until the reaction
was complete (monitored by 31P NMR). Inorganic salts were fil-
tered off through a Celite® pad and the residual solvent was re-
moved. The crude product was purified by silica gel flash
chromatography (eluent: pentane/ethyl acetate 7:3). White powder;
yield 95% (2.785 g, 2.736 mmol). 1H NMR (250 MHz, CDCl3): δ
= 6.84 (m, 24 H, H2,3), 4.68 (d, J = 2.4 Hz, 12 H, OCH2), 2.55 (t,
J = 2.4 Hz, 6 H, CϵCH) ppm. 13C{1H} NMR (75 MHz, CDCl3):
δ = 154.5 (C4), 144.8 (m, C1), 121.9 (C2), 115.5 (C3), 78.5 (CϵCH),
75.8 (CϵCH), 56.2 (OCH2) ppm. 31P{1H} NMR (101 MHz,
CDCl3): δ = 9.8 ppm. MS (FAB): m/z = 1017 [M]+. C54H42N3O12P3
(1017.9): calcd. C 63.72, H 4.16, N 4.13; found C 63.58, H 4.10, N
4.06.
31P{1H} NMR (162 MHz, [D ]THF): δ = 9.1 ppm. IR (neat): ν =
˜
8
3200 (br., OH) cm–1. C42H42N3O12P3 (873.7): calcd. C 57.74, H
4.85, N 4.81; found C 57.68, H 4.83, N 4.72.
Compound 2c: SOCl2 was added to a cooled (ice bath) round-bot-
tomed flask containing compound 2b (1 mmol) until complete dis-
solution. The reaction mixture was stirred overnight. Excess SOCl2
was removed under vacuum line with use of toluene as co-solvent,
and the crude product was used for the next step without further
purification. White powder; yield 93% (0.915 g, 0.93 mmol). 1H
Compound 6: A solution of propargylamine (30 mmol) and para-
formaldehyde (60 mmol, in H2O solution) in THF (20 mL) was in-
troduced into a two-necked flask fitted with a condenser. Dimethyl
hydrogen phosphonate (60 mmol) was then added dropwise, and
the reaction mixture was heated at 70 °C until the reaction was
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FULL PAPER
complete (monitored by 31P{1H} NMR). After evaporation of
THF, CHCl3 was added and the organic phase was washed with
NaOH (0.1 ), to extract excess dimethyl hydrogen phosphonate,
and dried with MgSO4. The crude residue was purified by silica gel
flash chromatography (CH2Cl2/MeOH 98:2). Pale yellow oil; yield
was evaporated, the residue was dissolved in CHCl3, the organic
phase was washed with water and brine and dried with MgSO4,
and the solvents were evaporated. The crude product was obtained
as a white foam and excess starting material was removed by flash
chromatography (eluent: CH2Cl2/MeOH 95:5). White foam; yield
1
1
76% (6.822 g, 22.80 mmol). H NMR (300 MHz, CDCl3) δ = 3.81
75% (0.225 g, 0.075 mmol). H NMR (400 MHz, CDCl3) δ = 7.86
(d, J = 10.5 Hz, 12 H, POCH3), 3.77 (d, J = 2.1 Hz, 2 H, CCH2N), (s, 6 H, CHtz), 6.75 (d, J = 8.8 Hz, 12 H, H2), 6.60 (d, J = 8.8 Hz,
3.14 (d, J = 10.5 Hz, 4 H, NCH2P), 2.30 (t, J = 2.1 Hz, 1 H, 12 H, H3), 4.76 (t, J = 5.0 Hz, 12 H, CH2CH2N), 4.28 (d, J =
CϵCH) ppm. 13C{1H} NMR (75 MHz, CDCl3): δ = 77.0 (CϵCH), 5.0 Hz, 12 H, OCH2CH2), 4.13 (s, 12 H, CCH2N), 3.72 (d, J =
74.6 (HCϵC), 52.5 (d, J = 6.7 Hz, POCH3), 48.8 (dd, J = 162.7, J
= 11.6 Hz, NCH2P), 45.5 (t, J = 8.6 Hz, CCH2N) ppm. 31P{1H} 13C{1H} NMR (100 MHz, CDCl3): δ = 154.9 (C4), 144.8 (m, C1),
NMR (121.5 MHz, CDCl3): δ = 25.9 ppm. MS (DCI/NH3): m/z =
143.3 (Ctz), 124.9 (CHtz), 121.8 (C2), 115.1 (C3), 66.9 (OCH2CH2),
300.0 [M + H]+. C9H19NO6P2 (299.2): calcd. C 36.13, H 6.40, N 52.8 (d, J = 6.9 Hz, POCH3), 51.1 (t, J = 8.1 Hz, CCH2N), 49.7
10.8 Hz, 72 H, POCH3), 3.21 (d, J = 10.4 Hz, 24 H, NCH2P) ppm.
4.68; found C 36.06, H 6.34, N 4.61.
(CH2CH2N), 49.0 (dd, J = 159.4, J = 9.5 Hz, NCH2P) ppm.
31P{1H} NMR (162 MHz, CDCl3): δ = 26.4 [PO(OCH3)2], 9.4
Compound 8.2: A solution of compound 8.1[25] (see Scheme 6,
14 mmol) and Et3N (84 mmol) in dry CH2Cl2 was cooled in an
ice bath. MsCl (70 mmol) was then added dropwise. The reaction
mixture was stirred at room temp. until the reaction was complete
(monitored by TLC and 31P{1H} NMR). NH4Cl (aq) was added
and the organic compounds were extracted with CH2Cl2. The or-
ganic phase was washed with brine and dried with MgSO4, and the
solvents were evaporated. The crude residue was purified by silica
gel flash chromatography (CH2Cl2/MeOH 99:1). Pale yellow oil;
(P0) ppm. MS (MALDI): m/z
102H162N27O48P15 (2999.1): calcd. C 40.85, H 5.44, N 12.61; found
C 40.72, H 5.42, N 12.48.
=
3021 [M
+
Na]+.
C
General Procedure for the Synthesis of the First and Second Genera-
tions of Dendrimers – 12-[G1] and 12-[G2]: Cs2CO3 (13.2 or
26.4 mmol) was added to a THF (dry, 20 mL) solution of den-
drimer 11-[G1] or 11-[G2] (0.5 mmol) and 4-(2-chloroethoxy)phenol
(6.6 or 13.2 mmol), and the reaction mixture was stirred at 40 °C
until the reaction was complete (12–36 h, monitored by 31P{1H}
NMR). Inorganic salts were filtered through a Celite® pad and the
residual solvent was removed. The crude product was purified by
silica gel flash chromatography in pentane/ethyl acetate (7:3).
1
yield 65% (3.615 g, 9.10 mmol). H NMR (300 MHz, CDCl3): δ =
4.37 (t, J = 6.4 Hz, 2 H, MsOCH2), 3.80 (d, J = 10.4 Hz, 12 H,
POCH3), 3.18 (d, J = 8.8 Hz, 4 H, NCH2P), 3.05 (s, 3 H, MsO),
2.95 (t, J = 6.4 Hz, 2 H, CH2CH2N), 1.95 (q, J = 6.4 Hz, 1 H,
CH2CH2CH2) ppm. 31P{1H} NMR (162 MHz, CDCl3):
δ =
Dendrimer 12-[G1]: White foam; yield 87% (0.602 g, 0.174 mmol).
26.7 ppm. C10H25NO9P2S (397.3): calcd. C 30.23, H 6.34, N 3.53;
found C 30.06, H 6.29, N 3.46.
1H NMR (300 MHz, CDCl3): δ = 7.63 (d, J = 8.4 Hz, 12 H, H3 ),
0
7.59 (s, 6 H, CH=N), 7.10 (d, J = 9.0 Hz, 24 H, H2 ), 7.00 (d, J =
1
8.4 Hz, 12 H, H2 ), 6.79 (d, J = 9.0 Hz, 24 H, H3 ), 4.12 (t, J =
0
1
5.1 Hz, 24 H, OCH2), 3.77 (t, J = 5.1 Hz, 24 H, CH2Cl) 3.24 (d, J
= 10.2 Hz, 18 H, CH3NP1) ppm. 13C{1H} NMR (75 MHz, CDCl3):
δ = 155.6 (C4 ), 151.2 (br. s, C1 ), 144.6 (d, J = 7.2 Hz, C1 ), 138.4
1
0
1
(d, J = 14.3 Hz, CH=N), 132.3 (C4 ), 128.3 (C3 ), 122.4 (d, J =
0
0
4.3 Hz, C2 ), 121.4 (C2 ), 115.4 (C3 ), 68.4 (OCH2), 41.9 (CH2Cl),
1
0
1
33.1 (d, J = 11.2 Hz, CH3NP1) ppm. 31P{1H} NMR (121.5 MHz,
CDCl3): δ = 64.2 (P1), 8.5 (P0) ppm. C144H144Cl12N15O30P9S6
(3461.4): calcd. C 49.97, H 4.19, N 6.07; found C 49.72, H 4.14, N
5.99.
Scheme 6. Synthesis of 8.
Dendrimer 12-[G2]: White foam; yield 79% (3.182 g, 0.395 mmol).
1H NMR (300 MHz, CDCl3): δ = 7.59 (m, 54 H, H3 , H3 , CH=N0,
Compound 8: NaN3 (4.5 mmol) was added to a solution of 8.2
(3 mmol) in H2O (10 mL), and the reaction mixture was stirred at
60 °C overnight. The aqueous phase was extracted with ethyl acet-
ate and the organic phase was washed with brine, dried with
MgSO4 and concentrated to give the azide derivative as a pale yel-
low oil; yield 90% (0.929 g, 2.70 mmol). 1H NMR (300 MHz,
CDCl3): δ = 3.80 (d, J = 10.8 Hz, 12 H, POCH3), 3.40 (t, J =
6.9 Hz, 2 H, N3CH2), 3.18 (d, J = 8.7 Hz, 4 H, NCH2P), 2.90 (t, J
0
1
CH=N1), 7.18 (d, J = 7.8 Hz, 24 H, H2 ), 7.08 (d, J = 8.7 Hz, 48
1
H, H2 ), 6.88 (d, J = 7.8 Hz, 12 H, H2 ), 6.78 (m, 48 H, H3 ), 4.10
2
0
2
(d, J = 5.7 Hz, 48 H, OCH2), 3.72 (d, J = 5.7 Hz, 48 H, CH2Cl),
3.24 (d, J = 10.2 Hz, 36 H, CH3NP2), 3.20 (d, J = 12.6 Hz, 18 H,
CH3NP1) ppm. 13C{1H} NMR (75 MHz, CDCl3): δ = 155.6 (C4 ),
2
151.2 (d, J = 7.0 Hz, C1 , C1 ), 144.5 (d, J = 7.0 Hz, C1 ), 139.1
1
0
2
(d, J = 12.9 Hz, CH=N1), 138.4 (d, J = 14.0 Hz, CH=N2), 132.4
(C4 ), 132.1 (C4 ), 128.3 (br. s, C3 , C3 ), 122.4 (d, J = 4.4 Hz, C2 ),
=
6.9 Hz, 2 H, CH2CH2N), 1.77 (q, J = 6.9 Hz, 2 H,
1
0
1
0
2
121.8 (d, J = 4.4 Hz, C2 ), 121.4 (br. s, C2 ), 115.4 (C3 ), 68.4
CH2CH2CH2) ppm. 13C{1H} NMR (100 MHz, CDCl3): δ = 53.9
(t, J = 6.6 Hz, CH2CH2N), 52.6 (d, J = 6.7 Hz, POCH3), 49.4 (dd,
1
0
2
(OCH2), 41.9 (CH2Cl), 33.1 (d, J = 12.5 Hz, CH3NP2), 32.9 (d, J
= 12.5 Hz, CH3NP1) ppm. 31P{1H} NMR (121.5 MHz, CDCl3):
δ = 64.3 (P2), 62.3 (P1), 8.5 (P0) ppm. C336H336Cl24N39O66P21S18
(8055.1): calcd. C 50.10, H 4.20, N 6.78; found C 50.00, H 4.17, N
6.66.
J
=
156.4,
J = 6.6 Hz, NCH2P), 49.0 (N3CH2CH2), 27.0
(CH2CH2CH2) ppm. 31P{1H} NMR (121.5 MHz, CDCl3): δ =
26.9 ppm. IR (neat): ν = 2095 (N ) cm–1. MS (DCI/NH ): m/z =
˜
3
3
345.1 [M + H]+. C9H22N4O6P2 (344.2): calcd. C 31.40, H 6.44, N
16.28; found C 31.17, H 6.41, N 16.15.
General Procedure for the Synthesis of Azide Derivatives 13-[G1] and
13-[G2]: NaN3 (6.6 or 13.2 mmol) was slowly added to a DMF (dry,
5 mL) solution of the corresponding chloride derivative 12-[G1] or
12-[G2] (0.5 mmol), and the reaction mixture was stirred and heated
overnight at 70 °C. The reaction mixture was then allowed to cool
at room temp., distilled water (5 mL) was added, and a white pre-
Dendrimer 10-[G0]: DIPEA (2 equiv. per azide) and CuI (0. 1 equiv.
per azide) were added to a degassed THF solution of dendrimer 3
(0.1 mmol) and alkyne 6 (1.1 equiv. per azide). The reaction mix-
ture was stirred at 40 °C until the reaction was complete (12–24 h,
monitored by IR, 31P{1H} NMR, 1H NMR). On completion, THF
2764
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© 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2010, 2759–2767

Design of Bisphosphonate-Terminated Dendrimers
cipitate appeared. The aqueous phase was decanted and the pre-
cipitate was dissolved in CH2Cl2. The organic phase was washed
with brine, dried with MgSO4 and concentrated to give the azide
derivative. If necessary, silica gel flash chromatography was per-
formed.
Dendrimer 14-[G2]: White foam; yield 79% (1.216 g, 0.079 mmol). 1H
NMR (400 MHz, CDCl3): δ = 7.82 (s, 24 H, CHtz), 7.61 (m, 54 H,
H3 , H3 , CH=N0, CH=N1), 7.20 (d, J = 8.8 Hz, 24 H, H2 ), 7.05
0
1
1
(d, J = 8.4 Hz, 48 H, H2 ), 6.91 (m, 12 H, H2 ), 6.76 (d, J = 8.4 Hz,
2
0
48 H, H3 ), 4.69 (m, 48 H, CH2CH2N), 4.26 (m, 48 H, OCH2CH2),
2
4.15 (s, 48 H, CCH2N), 3.75 (d, J = 11.2 Hz, 288 H, POCH3), 3.25
(d, J = 9.6 Hz, 54 H, CH3NP1, CH3NP2), 3.16 (d, J = 10.4 Hz, 96
Dendrimer 13-[G1]: White foam; yield 92% (1.628 g, 0.460 mmol).
1H NMR (300 MHz, CDCl3): δ = 7.64 (d, J = 8.4 Hz, 12 H, H3 ),
0
H, NCH2P) ppm. 13C{1H} NMR (75 MHz, CDCl3): δ = 155.2 (C4 ),
2
7.60 (s, 6 H, CH=N), 7.10 (d, J = 9.0 Hz, 24 H, H2 ), 7.01 (d, J =
1
151.2 (d, J = 7.0 Hz, C1 , C1 ), 144.5 (d, J = 7.1 Hz, C1 ), 143.3 (Ctz),
1
0
2
8.4 Hz, 12 H, H2 ), 6.79 (d, J = 9.0 Hz, 24 H, H3 ), 4.05 (t, J =
0
1
139.2 (br. s, CH=N1), 138.5 (d, J = 17.7 Hz, CH=N2), 132.4 (C4 ),
1
4.8 Hz, 24 H, OCH2), 3.54 (t, J = 4.8 Hz, 24 H, CH2N3) 3.25 (d,
J = 10.2 Hz, 18 H, CH3NP1) ppm. 13C{1H} NMR (75 MHz,
CDCl3): δ = 155.7 (C4 ), 151.2 (br. s, C1 ), 144.6 (d, J = 6.8 Hz,
132.1 (C4 ), 128.3 (C3 , C3 ), 124.7 (CHtz), 122.5 (d, J = 5.4 Hz, C2 ),
0
1
0
2
121.8 (C2 ), 121.3 (C2 ), 115.2 (C3 ), 66.7 (OCH2CH2), 53.8 (d, J =
1
0
2
1
0
7.0 Hz, POCH3), 51.1 (t, J = 8.0 Hz, CCH2N), 50.0 (CH2CH2N),
49.0 (dd, J = 159.4, J = 9.3 Hz, NCH2P), 33.1 (d, J = 12.3 Hz,
CH3NP2), 33.0 (d, J = 12.0 Hz, CH3NP1) ppm. 31P{1H} NMR
(121.5 MHz, CDCl3): δ = 64.5 (P2), 62.5 (P1), 26.4 [PO(OCH3)2], 8.4
(P0) ppm. C552H792N135O210P69S18 (15393.4): calcd. C 43.07, H 5.19,
N 12.28; found C 42.89, H 5.12, N 12.19.
C1 ), 138.4 (d, J = 14.2 Hz, CH=N), 132.3 (C4 ), 128.3 (C3 ), 122.4
1
0
0
(d, J = 4.5 Hz, C2 ), 121.4 (C2 ), 115.3 (C3 ), 67.4 (OCH2), 50.1
1
0
1
(CH2N3), 33.1 (d, J = 12.0 Hz, CH3NP1) ppm. 31P{1H} NMR
(121.5 MHz, CDCl ): δ = 64.3 (P ), 8.5 (P ) ppm. IR (neat): ν =
˜
3
1
0
2097 (N3) cm–1. C144H144N51O30P9S6 (3540.2): calcd. C 48.86, H
4.10, N 20.18; found C 48.61, H 4.01, N 20.09.
Dendrimer 15-[G0]: DIPEA (2 equiv. per azide) and CuI (0. 1 equiv.
per azide) were added to a degassed THF solution of dendrimer 5
(0.10 mmol) and azide 8 (1.1 equiv. per alkyne). The reaction
mixture was stirred at 40 °C until the reaction was complete (12–
24 h , monitored by IR, 31P{1H} NMR, 1H NMR). On completion,
THF was evaporated, the residue was dissolved in CHCl3, the or-
ganic phase was washed with water and brine and dried with
MgSO4, and the solvents were evaporated. The crude product was
obtained as a white foam and excess starting material was removed
by flash chromatography (eluent: CH2Cl2/MeOH 95:5). White
foam; yield 72% (0.222 g, 0.072 mmol). 1H NMR (300 MHz,
CDCl3): δ = 7.84 (s, 6 H, CHtz), 6.81 (m, 24 H, H2,3), 5.13 (s, 12
H, OCH2C), 4.48 (t, J = 7.2 Hz, 12 H, NtzCH2CH2CH2), 3.77 (d,
J = 10.5 Hz, 72 H, POCH3), 3.15 (d, J = 8.7 Hz, 24 H, NCH2P),
2.85 (t, J = 7.2 Hz, 12 H, CH2CH2NCH2), 2.06 (m, 12 H,
CH2CH2CH2) ppm. 13C{1H} NMR (100 MHz, CDCl3): δ = 155.2
(C4), 144.5 (C1), 143.5 (Ctz), 123.6 (CHtz), 121.8 (C2), 115.3 (C3),
62.3 (OCH2C), 53.7 (t, J = 7.3 Hz, CH2CH2N), 52.6 (d, J = 6.3 Hz,
Dendrimer 13-[G2]: White foam; yield 88% (3.614 g, 0.440 mol). 1H
NMR (300 MHz, CDCl3): δ = 7.63 (m, 42 H, H3 , CH=N0,
1
CH=N1), 7.56 (d, J = 8.4 Hz 12 H, H3 ), 7.19 (d, J = 7.8 Hz, 24
0
H, H2 ), 7.08 (d, J = 9.0 Hz, 48 H, H2 ), 6.88 (d, J = 8.4 Hz 12 H,
1
2
H2 ), 6.79 (m, 48 H, H3 ), 4.02 (d, J = 4.5 Hz, 48 H, OCH2), 3.51
0
2
(d, J = 4.5 Hz, 48 H, CH2N3), 3.24 (d, J = 10.4 Hz, 36 H,
CH3NP2), 3.20 (d, J = 11.2 Hz, 18 H, CH3NP1) ppm. 13C{1H}
NMR (75 MHz, CDCl3): δ = 155.6 (C4 ), 151.2 (d, J = 7.1 Hz,
2
C1 ,C1 ), 144.5 (d, J = 7.0 Hz, C1 ), 138.9 (br. s, CH=N1), 138.4
1
0
2
(d, J = 13.6 Hz, CH=N2), 132.4 (C4 ), 132.1 (C4 ), 128.3 (br. s, C3 ,
1
0
1
C3 ), 122.4 (d, J = 4.3 Hz, C2 ), 121.8 (C2 ), 121.4 (C2 ), 115.3
0
2
1
0
(C3 ), 67.3 (OCH2), 50.1 (CH2Cl), 33.1 (d, J = 12.5 Hz, CH3NP2),
2
32.9 (d, J = 12.9 Hz, CH3NP1) ppm. 31P{1H} NMR (121.5 MHz,
CDCl ): δ = 64.4 (P ), 62.3 (P ), 8.5 (P ) ppm. IR (neat): ν = 2095
˜
3
2
1
0
(N3) cm–1. C336H336N111O66P21S18 (8212.7): calcd. C 49.14, H 4.12,
N 18.93; found C 48.99, H 4.06, N 18.86.
General Procedure for the Synthesis of Dendrimers 14-[G1] and 14-
[G2] by “Click Chemistry”: DIPEA (2 equiv. per azide) and CuI
(0.1 equiv. per azide) were added to a degassed THF solution of
dendrimer 13-[G1] or 13-[G2] (0.1 mmol) and alkyne 6 (1.1 equiv.
per azide). The reaction mixture was stirred at 40 °C until the reac-
POCH3), 49.5 (dd,
(NtzCH2CH2), 28.4 (CH2CH2CH2) ppm. 31P{1H} NMR
(121.5 MHz, CDCl3): 26.8 [PO(OCH3)2], 9.7 (P0) ppm.
J = 157.0, J = 6.6 Hz, NCH2P), 47.7
δ
=
C108H174N27O48P15 (3083.3): calcd. C 42.07, H 5.69, N 12.27; found
C 42.03, H 5.62, N 12.22.
1
tion was complete (12–24 h, monitored by IR, 31P{1H} NMR, H
NMR). On completion, THF was evaporated, the residue was dis-
solved in CHCl3, the organic phase was washed with water and
brine and dried with MgSO4, and the solvents were evaporated.
The crude products were obtained as white foams and excess start-
ing material was removed by washing with THF.
General Procedure for the Synthesis of Dendrimers 16-[G1] and 16-
[G2]: Cs2CO3 (13.2 or 26.4 mmol) was added to a THF (dry, 20 mL)
solution of the dendrimer 11-[G1] or 11-[G2] (0.5 mmol) and 4-
(prop-2-yn-1-yloxy)phenol (6.6 or 13.2 mmol), and the reaction
mixture was stirred at 40 °C until the reaction was complete (12–
36 h, monitored by 31P{1H} NMR). Inorganic salts were filtered
through a Celite® pad and the residual solvent was removed. The
crude product was purified by silica gel flash chromatography in
pentane/ethyl acetate (7:3).
Dendrimer 14-[G1]: White foam; yield 85% (0.606 g, 0.085 mmol).
1H NMR (400 MHz, CDCl3): δ = 7.82 (s, 12 H, CHtz), 7.65 (d, J
= 8.5 Hz, 12 H, H3 ), 7.64 (s, 6 H, CH=N), 7.10 (d, J = 9.2 Hz, 24
0
H, H2 ), 7.01 (d, J = 8.5 Hz, 12 H, H2 ), 6.76 (d, J = 9.2 Hz, 24
1
0
H, H3 ), 4.71 (t, J = 4.8 Hz, 24 H, CH2CH2N), 4.27 (d, J = 4.8 Hz,
Dendrimer 16-[G1]: White foam; yield 91% (1.441 g, 0.455 mmol).
1
24 H, OCH2CH2), 4.15 (s, 24 H, CCH2N), 3.75 (d, J = 10.4 Hz,
144 H, POCH3), 3.23 (d, J = 10.0 Hz, 18 H, CH3NP1), 3.17 (d, J
1H NMR (300 MHz, CDCl3): δ = 7.62 (d, J = 8.7 Hz, 12 H, H3 ),
0
7.57 (s, 6 H, CH=N), 7.11 (d, J = 8.7 Hz, 24 H, H2 ), 7.02 (d, J =
1
= 10.4 Hz, 48 H, NCH2P) ppm. 13C{1H} NMR (75 MHz, CDCl3): 8.7 Hz, 12 H, H2 ), 6.86 (d, J = 8.7 Hz, 24 H, H3 ), 4.60 (d, J =
0
1
δ = 155.3 (C4 ), 151.3 (br. s, C1 ), 144.7 (d, J = 7.5 Hz, C1 ), 143.4
2.4 Hz, 24 H, OCH2C), 3.24 (d, J = 10.2 Hz, 18 H, CH3NP1), 2.50
(t, J = 2.4 Hz, 12 H, HCϵC) ppm. 13C{1H} NMR (75 MHz,
CDCl3): δ = 155.0 (C4 ), 151.2 (br. s, C1 ), 144.8 (d, J = 7.5 Hz,
1
0
1
(Ctz), 138.4 (br. s, CH=N), 132.3 (C4 ), 128.3 (C3 ), 124.7 (CHtz),
0
0
122.4 (d,
J =
3.8 Hz, C2 ), 121.4 (C2 ), 115.3 (C3 ), 66.7
1 0 1
1
0
(OCH2CH2), 52.8 (d, J = 7.1 Hz, POCH3), 51.0 (t, J = 8.3 Hz, C1 ), 138.4 (d, J = 13.6 Hz, CH=N), 132.3 (C4 ), 128.2 (C3 ), 122.3
1
0
0
CCH2N), 49.6 (CH2CH2N), 49.0 (dd, J = 159.4, J = 9.3 Hz, (d, J = 4.5 Hz, C2 ), 121.4 (C2 ), 115.7 (C3 ), 78.4 (HCϵC), 75.8
1
0
1
NCH2P), 33.1 (d, J = 11.7 Hz, CH3NP1) ppm. 31P{1H} NMR
(121.5 MHz, CDCl3): δ = 64.4 (P1), 26.5 [PO(OCH3)2], 8.4
(P0) ppm. C252H372N63O102P33S6 (7130.6): calcd. C 42.45, H 5.26,
N 12.38; found C 42.41, H 5.22, N 12.33.
(HCϵC), 56.2 (OCH2), 33.1 (d, J = 12.1 Hz, CH3NP1) ppm.
31P{1H} NMR (121.5 MHz, CDCl3): δ = 64.0 (P1), 8.4 (P0) ppm.
C156H132N15O30P9S6 (3168.0): calcd. C 59.15, H 4.20, N 6.63;
found C 59.10, H 4.10, N 6.55.
Eur. J. Org. Chem. 2010, 2759–2767
© 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
2765

E. Cavero, M. Zablocka, A.-M. Caminade, J. P. Majoral
FULL PAPER
Dendrimer 16-[G2]: White foam; yield 77% (2.875 g, 0.385 mmol).
P3N3Cl6 (2.88 mmol), and the mixture was stirred at 40 °C until
1H NMR (300 MHz, CDCl3): δ = 7.61 (m, 42 H, H3 , H3 , the reaction was complete (monitored by 31P{1H} NMR). Inor-
0
1
CH=N0), 7.54 (s, 12 H, CH=N1), 7.17 (d, J = 7.5 Hz, 24 H, H2 ), ganic salts were filtered through a Celite® pad and the residual
1
7.09 (m, 48 H, H2 ), 6.85 (m, 60 H, H3 , H2 ), 4.57 (d, J = 2.4 Hz, solution was evaporated. The crude product was purified by silica
2
2
0
48 H, OCH2C), 3.23 (d, J = 10.2 Hz, 36 H, CH3NP2), 3.18 (d, J = gel flash chromatography in CHCl3/MeOH (95:5); yield 60%
1
10.5 Hz, 18 H, CH3NP1), 2.48 (t, J = 2.4 Hz, 24 H, HCϵC) ppm. (1.358 g, 1.728 mmol). H NMR (300 MHz, [D6]acetone): δ = 7.35
13C{1H} NMR (75 MHz, CDCl3): δ = 154.9 (C4 ), 151.2 (d, J =
(d, J = 8.7 Hz, 4 H, H3), 7.28 (d, J = 8.7 Hz, 4 H, H3), 7.27 (d, J
= 8.7 Hz, 2 H, H3), 7.12 (d, J = 8.7 Hz, 4 H, H2), 6.92 (d, J =
2
7.1 Hz, C1 , C1 ), 144.7 (d, J = 7.1 Hz, C1 ), 139.0 (br. s, CH=N1),
1
0
2
138.4 (d, J = 13.6 Hz, CH=N2), 132.4 (C4 ), 132.1 (C4 ), 128.3 (br. 8.7 Hz, 4 H, H2), 6.87 (d, J = 8.7 Hz, 2 H, H2), 4.63 (s, 10 H,
1
0
s, C3 , C3 ), 122.4 (d, J = 4.4 Hz, C2 ), 121.8 (d, J = 4.4 Hz, C2 ),
CH2OH), 4.49 (br. s, 3 H, OH), 4.42 (m, 2 H, OH) ppm. 13C{1H}
1
0
2
1
121.5 (br. s, C2 ), 115.6 (C3 ), 78.4 (HCϵC), 75.8 (HCϵC), 56.2 NMR (75 MHz, [D6]acetone): δ = 149.2 (m, C1), 148.8 (d, J =
0
2
(OCH2), 33.1 (d, J = 12.5 Hz, CH3NP2), 32.9 (d, J = 12.4 Hz,
9.7 Hz, C1), 140.2 (d, J = 2.6 Hz, C4), 139.8 (C4), 139.6 (C4), 127.8
CH3NP1) ppm. 31P{1H} NMR (121.5 MHz, CDCl3): δ = 64.1 (P2), (C3), 127.75 (C3), 120.8 (m, C2), 120.5 (m, C2), 63.2 (CH2OH), 63.1
62.4 (P1), 8.6 (P0) ppm. C360H312N39O66P21S18 (7468.3): calcd. C (CH2OH) ppm. 31P{1H} NMR (121.5 MHz, [D6]acetone): δ = 22.5
57.90, H 4.21, N 7.31; found C 57.37, H 4.15, N 7.24.
(dd, J = 81.0, 85.0 Hz, P0), 7.2 (2ϫd, J = 81.0, 85.0 Hz, P0) ppm.
C35H35ClN3O10P3 (786.0): calcd. C 53.48, H 4.49, N 5.35; found C
53.42, H 4.48, N 5.33.
General Procedure for the Synthesis of Dendrimers 17-[G1] and 17-
[G2]: DIPEA (2 equiv. per alkyne) and CuI (0.1 equiv. per alkyne)
were added to a degassed THF solution of dendrimer 16-[G1] or
16-[G2] (0.1 mmol) and azide 8 (1.1 equiv. per alkyne). The reaction
mixture was stirred at 40 °C until the reaction was complete (12–
24 h , monitored by IR, 31P{1H} NMR, 1H NMR). On completion,
THF was evaporated, the residue was dissolved in CHCl3, the or-
ganic phase was washed with water and brine and dried with
MgSO4, and the solvents were evaporated. Crude products were
obtained as white foams and excess starting material was removed
by washing with THF.
Dendrimer 19: Cs2CO3 (1.65 mmol) was added to a THF (dry,
60 mL) solution of compound 18 (1.5 mmol) and 4-(prop-2-ynyl-
oxy)phenol (1.65 mmol), and the mixture was stirred at 40 °C until
the reaction was complete (monitored by 31P NMR). Inorganic
salts were filtered through a Celite® pad and the residual solvent
was removed. The crude product was purified by silica gel flash
chromatography (CHCl3/MeOH 95:5); yield 79% (1.064 g,
1
1.185 mmol). H NMR (300 MHz, [D6]acetone): δ = 7.28 (d, J =
8.4 Hz, 4 H, H3), 7.24 (d, J = 8.4 Hz, 6 H, H3), 6.9 (m, 10 H, H2),
6.89 (s, 4 H, H2Ј,3Ј), 4.79 (d, J = 2.4 Hz, 2 H, CH2CϵCH), 4.64
(m, 10 H, CH2OH), 4.45 (t, J = 5.7 Hz, 3 H, OH), 4.38 (t, J =
5.7 Hz, 2 H, OH), 3.11 (t, J = 2.4 Hz, 1 H, CϵCH) ppm. 13C{1H}
NMR (75 MHz, [D8]THF): δ = 154.9 (C4Ј), 149.5 (m, C1), 144.8
(m, C1Ј), 139.2 (C4), 139.1 (C4), 127.3 (C3), 127.2 (C3), 121.7 (m,
C2Ј), 120.5 (m, C2), 115.2 (C3Ј), 78.8 (CϵCH), 75.7 (CϵCH), 63.3
(CH2OH), 63.2 (CH2OH), 55.7 (CH2CϵCH) ppm. 31P{1H} NMR
(162 MHz, [D6]acetone): δ = 9.3 (m, P0) ppm. MS (DCI/NH3): m/z
= 898.2 [M + H]+. C44H42N3O12P3 (897.7): calcd. C 58.87, H 4.72,
N 4.68; found C 58.61, H 4.61, N 4.60.
Dendrimer 17-[G1]: White foam; yield 89% (0.650 g, 0.089 mmol).
1H NMR (250 MHz, CDCl3): δ = 7.78 (s, 12 H, CHtz), 7.65 (m, 18
H, CH=N, H3 ), 7.09 (d, J = 8.7 Hz, 24 H, H2 ), 7.01 (d, J =
0
1
9.0 Hz, 12 H, H2 ), 6.87 (d, J = 8.7 Hz, 24 H, H3 ), 5.10 (s, 24 H,
0
1
OCH2), 4.48 (t, J = 6.7 Hz, 24 H, CH2Ntz), 3.78 (d, J = 10.5 Hz,
144 H, POCH3), 3.24 (d, J = 10.3 Hz, 18 H, CH3NP1), 3.17 (d, J
= 9.0 Hz, 48 H, NCH2P), 2.85 (t, J = 6.7 Hz, 24 H, CH2CH2N),
2.07 (m, 24 H, CH2CH2CH2) ppm. 13C{1H} NMR (75 MHz,
CDCl3): δ = 155.8 (C4 ), 151.2 (br. s, C1 ), 144.4 (d, J = 7.5 Hz,
1
0
C1 ), 143.5 (Ctz), 138.7 (d, J = 13.5 Hz, CH=N), 132.2 (C4 ), 128.3
1
0
(C3 ), 123.5 (CHtz), 122.3 (d, J = 4.4 Hz, C2 ), 121.4 (C2 ), 115.4
0
1
0
Dendrimer 20-[G1]: DIPEA (2 equiv. per azide) and CuI (0.1 equiv.
per azide) were added to a degassed THF solution of dendrimer 4
(0.1 mmol) and alkyne 19 (1.1 equiv. per azide). The reaction mix-
ture was stirred at 40 °C until the reaction was complete
(C3 ), 62.3 (OCH2), 53.6 (t, J = 7.4 Hz, CH2CH2N), 52.6 (m,
1
POCH3), 49.5 (dd,
J
=
157.0,
J
=
6.8 Hz, NCH2P), 47.7
(NtzCH2CH2), 33.1 (d,
J
=
12.0 Hz, CH3NP1), 28.4
(CH2CH2CH2) ppm. 31P{1H} NMR (121.5 MHz, CDCl3): δ = 64.4
(P1), 26.8 [PO(OCH3)2], 8.5 (P0) ppm. C264H396N63O102P33S6
(7298.9): calcd. C 43.44, H 5.47, N 12.09; found C 43.22, H 5.41,
N 11.99.
1
(12–24 h , monitored by IR, 31P{1H} NMR, H NMR). On com-
pletion, THF was evaporated, the residue was dissolved in CHCl3,
the organic phase was washed with water and brine and dried with
MgSO4, and the solvents were evaporated. The crude product was
obtained as a white foam and excess starting material was removed
by flash chromatography (eluent: THF/MeOH 6:4). White foam;
Dendrimer 17-[G2]: White foam; yield 76% (1.195 g, 0.076 mmol).
1H NMR (300 MHz, CDCl3): δ = 7.78 (s, 24 H, CHtz), 7.62 (m, 54
H, H3 , H3 , CH=N0, CH=N1), 7.17 (m, 24 H, H2 ), 7.07 (m, 48
1
yield 62% (0.397 g, 0.062 mmol). H NMR (300 MHz, [D8]THF):
0
1
1
δ = 8.1 (s, 6 H, CHtz), 7.19 (d, J = 8.4 Hz, 24 H, H3 ), 7.18 (d, J
H, H2 ), 6.87 (m, 60 H, H3 , H2 ), 5.07 (s, 48 H, OCH2), 4.46 (m,
1
2
2
0
= 8.4 Hz, 36 H, H3 ), 7.14 (d, J = 8.8 Hz, 12 H, H3 ), 6.93–6.86
48 H, CH2Ntz), 3.76 (m, 288 H, POCH3), 3.23–3.12 (m, 150 H,
CH3NP1, CH3NP2, NCH2P), 2.83 (m, 48 H, CH2CH2N), 2.04 (m,
48 H, CH2CH2CH2) ppm. 13C{1H} NMR (75 MHz, CDCl3): δ =
155.7 (C4 ), 151.2 (d, J = 7.1 Hz, C1 , C1 ), 144.5 (d, J = 7.1 Hz,
1
0
(m, 72 H, H2 , H2 ), 6.80 (m, 24 H, H2Ј,3Ј0), 5.57 (s, 12 H, CH2Ntz),
1
0
5.17 (s, 12 H, CH2O), 4.64 (m, 12 H, OH), 4.58 (d, J = 5.6 Hz, 24
H, CH2OH), 4.57 (d, J = 5.6 Hz, 36 H, CH2OH), 4.57 (m, 18 H,
OH) ppm. 13C{1H} NMR (75 MHz, [D8]THF): δ = 155.6 (C4Ј0),
150.3 (m, C1 ), 149.5 (m, C1 ), 144.4 (m, C1Ј0), 144.0 (Ctz), 139.1
2
1
0
C1 ), 143.5 (Ctz), 139.0 (br. s, CH=N1, CH=N2), 132.4 (C4 ), 132.1
2
1
(C4 ), 128.4 (C3 , C3 ), 123.6 (CHtz), 122.4 (d, J = 4.4 Hz, C2 ),
0
1
0
1
0
2
(C4 ), 132.8 (C4 ), 129.3 (C3 ), 127.4 (C3 ), 127.3 (C3 ), 124.0
121.8 (d, J = 4.4 Hz, C2 ), 121.4 (br. s, C2 ), 115.4 (C3 ), 62.3
1
0
0
1
1
1
0
2
(CHtz), 121.8 (C2Ј0), 121.1 (C2 ), 120.5 (C2 ), 115.1 (C3Ј0), 63.3
(OCH2), 53.7 (t, J = 7.7 Hz, CH2NCH2), 52.7 (m, POCH3), 49.5
(dd, J = 157.0, J = 6.8 Hz, NCH2P), 47.7 (NtzCH2CH2), 33.2 (d, J
= 12.0 Hz, CH3NP2, CH3NP1), 28.5 (CH2CH2CH2) ppm. 31P{1H}
0
1
(CH2OH), 63.27 (CH2OH), 61.9 (CH2O), 52.6 (CH2Ntz) ppm.
31P{1H} NMR (162 MHz, [D6]acetone): δ = 8.9 (s, P0), 9.3 (m,
P1) ppm. MS (MALDI): m/z
=
6432 [M
+
Na]+.
NMR (121.5 MHz, CDCl3):
δ = 64.5 (P2), 62.5 (P1), 26.8
C306H288N39O78P21 (6410.3): calcd. C 57.34, H 4.53, N 8.52; found
C 57.12, H 4.51, N 8.41.
[PO(OCH3)2], 8.4 (P0) ppm. C576H840N135O210P69S18 (15730.1):
calcd. C 43.98, H 5.38, N 12.02; found C 43.71, H 5.22, N 11.85.
Dendrimer 18: Cs2CO3 (31.68 mmol) was added to a THF (dry,
60 mL) solution of 4-hydroxybenzyl alcohol (15.84 mmol) and
Dendrimer 21-[G1]: SOCl2 was added to a cooled (ice bath) round-
bottomed flask containing dendrimer 20-[G1] (0.04 mmol) until
2766
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© 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2010, 2759–2767

Design of Bisphosphonate-Terminated Dendrimers
complete dissolution. The reaction mixture was stirred overnight.
Excess SOCl2 was removed under a vacuum line with use of toluene
as co-solvent and the crude product was used for the next step
without further purification. White foam. 1H NMR (300 MHz,
Acknowledgments
We thank the Centre National de la Recherche Scientifique
(CNRS). M. Z., A.-M. C., and J. P. M. thank the COST TD 0802
Action, and E.C. thanks the Fundacion Ramon Areces for finan-
cial support.
CDCl3): δ = 7.70 (s, 6 H, CHtz) 7.20 (2ϫd, J = 8.4 Hz, 60 H, H3 ),
1
7.04 (d, J = 8.4 Hz, 12 H, H3 ), 6.89 (d, J = 8.4 Hz, 60 H, H2 ),
0
1
6.85–6.75 (m, 36 H, H2 , H2Ј,3Ј0), 5.45 (s, 12 H, CH2Ntz), 5.11 (s,
0
12 H, CH2O), 4.55, 4.54, 4.52 (3ϫs, 60 H, CH2Cl) ppm. 13C{1H}
NMR (75 MHz, CDCl3): δ = 155.4 (C4Ј0), 150.3 (m, C1 , C1 ),
[1] S. H. Medina, M. E. H. El-Sayed, Chem. Rev. 2009, 109, 3141–
3157.
0
1
144.3 (m, C1Ј0, Ctz), 134.2 (C4 ), 131.8 (C4 ), 129.9 (C3 ), 129.3 [2] C. C. Lee, J. A. MacKay, J. M. J. Fréchet, F. C. Szoka, Nat. Bi-
1
0
1
(C3 ), 123.5 (CtzH), 122.0 (C2Ј0), 121.3 (C2 ), 121.1 (C2 ), 115.1
otechnol. 2006, 23, 1517–1526.
0
0
1
(C3Ј0), 62.2 (CH2O), 53.3 (CH2Ntz), 45.7, 45.6 (2ϫs, CH2Cl) ppm.
31P{1H} NMR (121.5 MHz, CDCl3): δ = 8.3 (s, P0), 8.8 (m,
P1) ppm.
[3] E. R. Gilies, J. M. J. Fréchet, Drug Deliv. Today 2005, 10, 35–
43.
[4] U. Boas, P. M. H. Heegaard, Chem. Soc. Rev. 2004, 33, 43–63.
[5] Y. Cheng, J. Wang, T. Rao, X. He, T. Xu, Front. Biosci. 2008,
13, 1447–1471.
Dendrimer 22-[G1]: NaN3 (1.6 mmol) was slowly added to a DMF
(dry, 5 mL) solution of dendrimer 21-[G1] (0.05 mmol), and the re-
action mixture was stirred and heated overnight at 70 °C. The reac-
tion mixture was then allowed to cool at room temp., distilled water
(5 mL) was added, and a white precipitate appeared. The aqueous
phase was decanted and the precipitate was dissolved in CH2Cl2.
The organic phase was then washed with brine, dried with MgSO4
and concentrated to give the azide derivative. White foam; yield
87% (0.311 g, 0.0435 mmol). 1H NMR (300 MHz, CDCl3): δ =
[6] M. J. Cloninger, Curr. Opin. Chem. Biol. 2002, 6, 742–748.
[7] O. Rolland, C.-O. Turrin, A.-M. Caminade, J.-P. Majoral, New
J. Chem. 2009, 33, 1809–1824.
[8] A. S. Chauhan, P. Diwan, N. K. Jain, D. A. Tomalia, Bioma-
cromolecules 2009, 10, 1195–1202.
[9] S. Fruchon, M. Poupot, L. Martinet, C.-O. Turrin, J.-P. Majo-
ral, J.-J. Fournié, A.-M. Caminade, R. Poupot, J. Leukocyte
Biol. 2009, 85, 553–562.
[10] P. Marchand, L. Griffe, M. Poupot, C.-O. Turrin, G. Bacquet,
J.-J. Fournié, J.-P. Majoral, R. Poupot, A.-M. Caminade, Bio-
org. Med. Chem. Lett. 2009, 19, 3963–3966.
[11] M. Poupot, L. Griffe, P. Marchand, A. Maraval, O. Rolland,
L. Martinet, F.-E. LЈFaqihi-Olive, C.-O. Turrin, A.-M. Camin-
ade, J.-J. Fournié, J.-P. Majoral, R. Poupot, FASEB J. 2006, 20,
2339–2351.
[12] L. Griffe, M. Poupot, A. Maraval, C.-O. Turrin, O. Rolland, P.
Métivier, G. Bacquet, J.-J. Fournié, A.-M. Caminade, R. Pou-
pot, J.-P. Majoral, Angew. Chem. Int. Ed. 2007, 46, 2523–2526.
[13] For a review on click chemistry and dendrimers see: G. Franc,
A. Kakkar, Chem. Commun. 2008, 42, 5267–5276.
[14] P. Wu, A. K. Feldman, A. K. Nugent, C. J. Hawker, A. Scheel,
B. Voit, J. Pyun, J. M. J. Fréchet, K. B. Sharpless, V. V. Fokin,
Angew. Chem. Int. Ed. 2004, 43, 3928–3932.
7.70 (s, 6 H, CHtz), 7.14 (m, 60 H, H3 ), 7.04 (d, J = 9.0 Hz, 12 H,
1
H3 ), 6.98 (m, 72 H, H2Ј0, H2 ), 6.86 (d, J = 9.0 Hz, 12 H, H3Ј0),
0
1
6.77 (d, J = 9.0 Hz, 12 H, H2 ), 5.45 (s, 12 H, CH2Ntz), 5.10 (s, 12
0
H, CH2O), 4.31, 4.30, 4.29 (3ϫs, 60 H, CH2N3) ppm. 13C{1H}
NMR (75 MHz, CDCl3): δ = 155.5 (C4Ј0), 150.3 (m, C1 , C1 ),
0
1
144.3 (m, C1Ј0), 144.2 (Ctz), 132.3 (C4 ), 131.8 (C4 ), 129.4 (C3 ),
1
0
1
129.2 (C3 ), 123.3 (CtzH), 121.9 (C2Ј0), 121.2 (C2 , C2 ), 115.4
0
0
1
(C3Ј0), 62.3 (CH2O), 54.1, 54.05 (2ϫs, CH2N3), 53.2
(CH2Ntz) ppm. 31P{1H} NMR (121.5 MHz, CDCl3): δ = 8.3 (s, P0),
8.6 (m, P1) ppm. IR (neat):
ν
˜
=
2092 (N3) cm–1.
C306H258N129O48P21 (7160.7): calcd. C 51.33, H 3.63, N 25.23;
found C 51.11, H 3.57, N 25.12.
[15] M. J. Joralemon, R. K. O’Reilly, J. B. Matson, A. K. Nugent,
C. J. Hawker, K. L. Wooley, Macromolecules 2005, 38, 5436–
5443. Addition/Correction: M. J. Joralemon, R. K. O’Reilly,
J. B. Matson, A. K. Nugent, C. J. Hawker, K. L. Wooley, Mac-
romolecules 2006, 39, 900.
[16] N. Launay, M. Slany, A.-M. Caminade, J.-P. Majoral, J. Org.
Chem. 1996, 61, 3799–3805.
[17] M.-L. Lartigue, M. Slany, A.-M. Caminade, J.-P. Majoral,
Chem. Eur. J. 1996, 2, 1417–1426.
[18] J.-C. Blais, C.-O. Turrin, A.-M. Caminade, J.-P. Majoral, Anal.
Chem. 2000, 72, 5097–5105.
Dendrimer 23-[G1]: DIPEA (2 equiv. per azide) and CuI (0.1 equiv.
per azide) were added to a degassed THF solution of dendrimer 22-
[G1] (0.05 mmol) and alkyne 6 (1.1 equiv. per azide). The reaction
mixture was stirred at 40 °C until the reaction was complete (12–
24 h, monitored by IR, 31P{1H} NMR, 1H NMR). On completion,
THF was evaporated, and the crude residue was directly purified
by silica gel flash chromatography; yield 88% (0.710 g,
0.044 mmol). 1H NMR (500 MHz, CDCl3): δ = 7.99 (s, 6 H,
CHtz0), 7.78 (s, 18 H, CHtz1), 7.71 (s, 12 H, CHtz1), 7.15 (m, 12 H,
[19] C. Galliot, C. Larré, A.-M. Caminade, J.-P. Majoral, Science
1997, 277, 1981–1984.
H3 ), 7.08 (m, 60 H, H3 ), 6.91 (m, 12 H, H2 ), 6.86 (m, 72 H, H2Ј
0
,
,
0
1
0
H2 ), 6.77 (m, 12 H, H3Ј0), 5.57, 5.54, 5.51 (3ϫs, 72 H, CH2Ntz0
1
[20] S. Merino, L. Brauge, A.-M. Caminade, J.-P. Majoral, D. Ta-
ton, Y. Gnanou, Chem. Eur. J. 2001, 7, 3095–3105.
[21] V. Cadierno, M. Zablocka, B. Donnadieu, A. Igau, J.-P. Majo-
ral, A. Skowronska, Chem. Eur. J. 2000, 6, 345–352.
[22] J.-P. Majoral, M. Zablocka, New J. Chem. 2005, 29, 32–41.
CH2Ntz1), 5.13 (s, 12 H, CH2O), 4.14 (s, 36 H, Ctz1CH2N), 4.10 (s,
18 H, Ctz1CH2N), 3.75–3.68 (m, 360 H, POCH3), 3.16, 3.13 (2ϫd,
J = 10.0 Hz, 120 H, NCH2P) ppm. 13C{1H} NMR (75 MHz,
CDCl3): δ = 155.7 (C4Ј0), 150.4 (m, C1 , C1 ), 144.2 (m, C1Ј0), 143.9,
0
1
143.8 (Ctz), 132.1 (m, C4 , C4 ), 129.5 (C3 ), 129.2, 129.1 (2ϫs, [23] E. Martinez-Ferrero, G. Franc, S. Mazères, C.-O. Turrin, A.-
1
0
0
C3 ), 124.0, 123.9 (2ϫs, CHtz), 121.9 (C2Ј0), 121.2 (2ϫs, C2 , C2 ),
M. Caminade, J.-P. Majoral, C. Sanchez, Chem. Eur. J. 2008,
14, 7658–7669.
[24] M. Srinivasan, S. Sankararaman, H. Hopf, I. Dix, P. G. Jones,
J. Org. Chem. 2001, 66, 4299–4303.
[25] K. Chougrani, B. Boutevin, G. David, G. Boutevin, Eur. Polym.
J. 2008, 44, 1771–1781.
1
0
1
115.5 (C3Ј0), 62.3 (CH2O), 53.2 (CH2Ntz), 52.8 (d, J = 6.9 Hz,
POCH3), 51.2 (t, J = 8.8 Hz, CtzCH2N), 49.0 (dd, J = 159.5, J =
9.2 Hz, NCH2P) ppm. 31P{1H} NMR (121.5 MHz, CDCl3): δ =
26.4 [s, PO(OCH3)2] 8.4 (s, P0), 8.5 (m, P1) ppm.
C576H828N159O228P81 (16136.6): calcd. C 42.87, H 5.17, N 13.80;
found C 42.16, H 5.06, N 13.59.
Received: November 10, 2009
Published Online: March 25, 2010
Eur. J. Org. Chem. 2010, 2759–2767
© 2010 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
2767