Journal of Nuclear and Radiochemical Sciences, Vol. 6, No. 3, pp. 173-176, 2005

ArticlesThe Facettes of [99TcCl3 (CO) 3] 2– Chemistry and Its Application to Life Science
Roger Alberto*
Institute of Inorganic Chemistry, University of Zürich, Winterthurerstr. 190, CH-8057 Zürich, Switzerland
Received: July 31, 2005; In Final Form: September 30, 2005
This article gives a brief summary about the current status of radiochemistry and radiopharmaceutical chemistry
as related to the fac-[99mTc(OH2)3(CO)3]+ and fac-[99Tc(OH2)3(CO)3]+ synthon. Since the synthesis and availabilityof synthons or direct precursors for the labeling of biomolecules (and for the exploration of fundamental inorganicand organometallic chemistry) is a crucial prerequisite for developing new radiopharmaceuticals, some consideration about composition and characteristics of such complexes are discussed at the beginning and their suitability
is discussed. Beside widely investigated basic and applied Tc chemistry in the oxidation state (V) with the[Tc=O]3+ or [Tc≡N] 2+ moieties, the aforementioned [99(m)Tc(CO)3]+ and [99Tc(CO)3]+ core are new members in thisgroup. Although its pressureless synthesis from organic solvents has been described about eight years ago, it is
only recently that exploitation of its radiopharmaceutical potential could reasonably start owing to the availabilityof a routinely applicable kit. We will summarize basic coordination chemistry of the [ 99 (m) Tc(CO) 3 ] + and[99Tc(CO)3] + moieties as relevant for radiopharmaceutical purposes, focusing on some fundamental reactions in
the context of low valent Tc chemistry. Due to its special properties, new research directions possible only throughits very special physico-chemical properties will be emphasized as well.

Synthons. The importance of stable but reactive precursors,
so called synthons, is very well recognized in synthetic inorganic chemistry. A synonym for synthon is “building block”,
thus, kind of a brick stone for constructing different structures.
The situation in chemistry is related to architecture and one of
the most important and common building blocks in technetiumchemistry is probably [TcOCl4]− from which the preparation ofmost novel complexes started.1−4 The synthon [TcOCl4]− is well
suited for chemistry in organic solvents, but, switching to aqueous solutions makes it less versatile due to its tendency tohydrolyze and to disproportionate. Real technetium synthonsstable in water are rare but highly important since the driving
force for research in technetium chemistry is usually the application in radiopharmacy. Suitable properties of such an aqueous
synthon are easily defined; i) for well characterized composition, ii) of sufficient stability towards hydrolysis or redox
decomposition, iii) exchangeable ligands and optionally iv)
forming robust complexes. Typically, aquo-ions of the lanthanides and other triply positive metal-cations fit well in thisscheme i) – iii) and are therefore most widely explored in radiopharmaceutical chemistry.5 The situation is different in technetium chemistry, where no real aquo-ions exist. The typicalprecursors in technetium chemistry comprise the [Tc = O]3+ andto a lower extent the [Tc ≡ N]2+ core. Despite being very useful,they are not real synthons since they often lack accurate characterization, long term stability or require the presence of additional organic solvents. They can however being stabilized insitu with strong multidentate chelators based on pure or mixed
sets of nitrogen, oxygen and sulfur donors or phosphorous
donors the number or combination of which is often not very
flexible. It is suggested to ask the question at this point whatkind of synthon one would ideally like to have for radiopharmaceutical research and development. This is not easily
answered and depends strongly on the particular problem.
Such compounds probably would have exchangeable water
ligands in common, independent of oxidation state or electronic configuration. The synthesis of [99mTc(OH2)3(CO)3]+ as*Corresponding author. E-mail: ARIEL@ACI.UNIZH.CH. FAX:
+ 41 1 635 68 03.

-[TcO4]

i)

2-

Cl
OC

OC

Tc

Cl

Cl

CO

ii)

OH2
OC

OC

Tc

OH2

OH2

CO

iii)
99(m)

Scheme 1. Synthesis of [TcCl3(CO)3] 2− and [99(m)Tc(OH2)3(CO)3]+99(m)−from [ TcO4] : i) H3B·THF, THF 1 atm CO; ii) H2O; iii) Na[H3BCO2H],
0.9% NaCl, 20 min, 95 ˚C (Isolink Kit®).

shown in Scheme 1 approaches an ideal synthon in that it fulfills, from a chemical point of view, most of the requirements
outlined above.6−9 Throughout the following text, the formulation “[99(m)Tc(OH2)3(CO)3]+” refers to experiments which havebeen performed with long-lived 99Tc and with short-lived 99mTc.
The [99mTc(OH2)3(CO)3]+ synthon is synthesized from water,comprises exchangeable water ligands and is stable even on
the no carrier added 99mTc level over extended time periods.
Depending on the view point, the three CO ligands might beconsidered a disadvantage since they provide lipophilicity andare relatively large in comparison to a potential water ligand.
However, it is an advantage that they occupy three coordinationsites irreversibly and potential ligands must have a maximumof three donors. Ligands with three donors only (or less) might
be small and even very small ligands can be designed such as
cyclopentadienyl systems (see later in the text).10 The reactivity
and coordination chemistry of [99(m)Tc(OH2)3(CO)3]+ with many
types of mono- bi- and tridentate ligands have been studied indetail but still new surprising features and research directions
emerge. Scheme 2 gives an overview on the most importantand common ligands for “stand alone chemistry” or suitablefor being attached to targeting vectors.11−18
Cyclopentadienyl Chemistry. The example of cyclopentadienyl (Cp) ligands shall be discussed in more detail and in
front of a general consideration. While reactions with most of
the ligands depicted in Scheme 2 are of the “Werner type”, Cp
is an organometallic system which chemistry seems not reallyto be related to water as a solvent. Cyclopentadiene is not stable or soluble in water but the corresponding ligand with anacetyl group (acp) is. Deprotonation occurs easily and at phys-© 2005 The Japan Society of Nuclear and Radiochemical Sciences
Published on Web 12/28/2005

174 J. Nucl. Radiochem. Sci., Vol. 6, No. 3, 2005

Alberto

H
N

HN

COOH

CN

N

OC

N

COOH

H2N

N

R

NH2

N

S

COOH

S

S

S

NH2

N

N
H

NH2

Scheme 2. A selection of divers ligands suitable for strong coordination to the fac-[Tc(CO)3]+ moiety in aqueous solution.

O1
C8

C4
C5

C6

C9

C3
C7

C10
C13

C2

C1

Tc1

C16

O16

C11
C12
O2

C14
O14

CO

O

HN

Tc

OC

OC

NH2
CO

N

NLS

CO
CO
B

C

Scheme 3. Representation of intercalating complexes based on thefac-[Tc(CO)3] + moiety. The bottom scheme shows the trifunctional
system, (A) intercalator, (B) ligand and (C) nucleus targeting portion.

H

N

NH2

Tc

A

NH2

O

N
H

COOH

COOH
H
B
N
N

CO

HN
OC

N

N

Tc

NH

O

NH

COOH

COOH

COOH

HN

O

NH

C15
O15

Figure 1. ORTEP presentation of [(H 3COC6H4 COC5H4)Tc(CO) 3].
Ellipsoids drawn with 50% probability.

iological pH about 30% is present in the anionic acetyl-cyclopentadienyl form. The demand for performing Cp chemistry inwater emerges from the need of producing small complexes aslabels, in particular for targeting brain receptor ligands. We
found that the synthon [99(m)Tc(OH2)3(CO)3]+ reacts with acetylcyclopentadienyl (acp) in water to form the corresponding
piano-stool complexes [(RCp)Tc(CO)3] in quantitative yield.19
This kind of research is nowadays categorized as bioorganometallic chemistry, a new and rapidly growing field of research.20
The problems arising from the compulsory limitation of using
water as a solvent might open the pathway to aqueous Cp chemistry with other transition metals as well. Technetium chemistry
does not only following synthetic pathways from e.g. rhenium
but is initiating new synthetic research directions. This situationis very uncommon and rarely encountered in the past. One of
the few exceptions is possibly the synthesis of the most important radiopharmaceutical [Tc(CN-R) 6 ] + as introduced by
Davison and coworkers.21 An ORTEP presentation of a Cp complex of technetium is given in Figure 1.22, 23
Intercalators. A further and rather provocative example inthe context of new possibilities is the combination of 99mTc withintercalating cell nucleus seeking molecules such as acridine,
acridine orange of pyrene. A less known characteristic featureof 99mTc is its possibility to emit about 2 Auger and KosterCronig electrons per decay (and electrons resulting from internal conversion).24 Auger electron emitters such as 111In are
under relatively strong research as potential radiotherapeutic
compounds. Although Auger electrons have a low energy, they
have a high LET and if a corresponding radionuclide decays in
the intimate vicinity of DNA, it can induce lethal double strand
breaks.25, 26 The challenge with this objective in mind is highsince a corresponding compound should target specific cancer
cell, being internalized, overcome the cell and nucleus walland finally bind to DNA by groove binding or intercalation. If
outside the nucleus, the Auger electrons have hardly a sufficient range to damage the cell’s DNA or other essential cellcompartments. Proof of principal has been given with 111In but
the approach is in general still considered to be “academic”
without naming principal reasons for the exclusion of this possibility. Clearly, a sufficient number of radionuclides must betransported to and deposited in the target but the same is also
true for (completely unspecific) chemotoxic drugs such as cisplatin. Using [99mTc(OH2)3(CO)3]+ as the building block, compounds can be designed which are cationic for better interaction
with the negatively charged DNA backbone and carrying anintercalator and a nucleus targeting agents such as nuclear
localizing sequence (NLS) peptides the same time. The principal design of such trifunctional molecules is depicted in
Scheme 3.
As could be shown, the [ 99mTc(OH 2) 3 (CO) 3] + based complexes with a pendant pyrene have in vitro the ability of inducing an activity dependent amount of single and double strand
breaks. Combined to the NLS peptides, these radiobioconjugates reduced the cell viability to a significant extent. Cells
exposed to equal amounts of [99mTcO 4] − or to the “cold” rhenium surrogates had no effect at all.27 The synthesis of the trifunctional molecule, ligand plus intercalator plus nucleus
targeting molecule, is relatively demanding and of low flexibility. We have altered the approach by selecting a more flexible[2+1] mixed ligand strategy in which the targeting function is
attached to a monodentate ligand and the intercalator to a bidentate chelator or vice versa.28 This will enable more convenient(fine) tuning of the systems or to select different nucleus seeking agents without the need of repeating complicated multistep
organic syntheses every time. The topic of applying 99mTc fortherapeutic purposes is the subject of controversial discussions
for reasons as outlined above. Such reasons might be true or
not but it should be emphasized that the same approach or strategy can also be verified with 188/186Re being considered to beamong the most important therapeutic radionuclides.
The advantage of using [99mTc(OH 2) 3 (CO) 3] + as a building
block is evident in this example. A complicated trifunctional
biomolecule simply reacts with this synthon under mild conditions. The concept could also be realized with any of the other
Tc(V) based precursors but would probably be more complicated, leading to byproducts due to the presence of a multicomponent system.
Vitamin B12. So far, ligands coordinated to the fac-[Tc(CO)3]+
moiety were bi- or tridentate for reasons outlined elsewhere.16
It has been described that even monodentate ligands can replacewater and form highly inert complexes. Typical examples arearomatic amines such as imidazole or pyridine but also phosphines and thioethers are appropriate groups. The coordination
to monodentate ligands can for instance be used for the directlabeling of biomolecules bearing corresponding coordinating
functionalities. A typical example in this respect are the imidazole in the side chains of histidine which have repeatedly

The Facettes of [99TcCl3 (CO) 3] 2− Chemistry

J. Nucl. Radiochem. Sci., Vol. 6, No. 3, 2005 175

Re
N
C

Figure 3. ORTEP presentation of [99Tc(HOCH3)(9-MeG) 2 (CO)3]+.

Figure 2. ORTEP presentation of vitamin B12 with [Co]-CN-[Re]bridging cyanide and coordinated [Re(NO)(CO)3] (NO = N,N-dimethylglycine).

been described for labeling with [ 99mTc(OH 2 ) 3 (CO) 3] +. 29, 30
Coordinated cyanide is one of the oldest “ligand” known ininorganic chemistry for forming bridges [M]-CN-[M’] between
two metal centers. Since cyanides are not common in biological systems, the application of those for labeling purposes is
unknown. Vitamin B12 is the exception and comprises in its
basic structure a cyanide coordinated to the central Co(III)cation in vitamin B12. We found that this cyanide is very
prone to bind to [99mTc(OH2)3(CO)3]+, thereby forming a complex as depicted in Figure 2.31
Although the cyanide in this example is a bridging monodentate ligand only, the coordination is very strong. The technetium
complex bound to it is only very slowly released even in thepresence of serum proteins. Two coordination sites remain in
principle occupied by water or chloride and might therefore
interact with other competing donors. The stability in serum
proves the contrast but these two sites can also be occupied withan anionic bidentate ligand to shield the Tc(I) center from further interactions. Since many bidentate ligands can be considered for this purpose, a (fine) tuning of the biological properties
becomes possible without changing the overall topology of theB12 derivative. Clearly, with an asymmetric bidentate ligand
two diastereomers are formed which have to be investigatedafter separation. It is still remarkable that the two diastereomeric forms, once separated from each other, do not interconvert
at all proving the high kinetic or thermodynamic stability of the[Co]-CN-[Tc](Re) bond.31 Since the important transport proteins for B12 in humans bind essentially to the lower hemisphereof B12, the introduction of a metal complex at the upper should
not interfere to a large extent with this recognition, an important
fact that is confirmed by preliminary biological studies.
Again, it can be emphasized here that the experimental findings with technetium inspired research with other transition
metals as well. It is surprising that no studies with B12 andbinding to other metals have been presented before. We foundthat other metal complexes or even building blocks such as different Pt(II) complexes readily bind to cyanide, a fact which is
not surprising for an inorganic chemist but still has not been
attempted before.32 The far goal of such a strategy in the context of technetium or other metals is the use of vitamin B12 asa Trojan horse for radio- and/or chemotoxic compounds.

Nucleobases. As a last example, the reactivity towards
guanine should briefly be mentioned. As the reaction withvitamin B12 is a good example for a stable monodentate ligand,[99mTc(OH2)3 (CO)3]+ and its rhenium analogue are also prone
for coordination to the nucleobase guanine. N7 in guanine is
the preferred target of cisplatin and its irreversible coordination
to two adjacent guanines in DNA is believed to be the reason
for its chemotoxic action.33−35 Octahedral complexes are notgenerally considered to copy this reactivity since the relatively
bulky nucleobases in isolated form or integrated in DNA mightsterically interfere with the other ligands. We have studied thereaction of [99mTc(OH 2) 3 (CO) 3] + with 9-methyl guanine (9MeG) and 7-methyl guanine (7-MeG) and found that two guanines coordinate relatively rapid to Tc(I).36 An example of thecomplex [99Tc(HOCH3)(9-MeG)2 (CO)3]+ is given in Figure 3.
The compound is stable in serum but its formation on the no carrier added level with 99mTc slow. Still, since [99mTc(OH2)3(CO)3]+
is obviously able to adopt two guanines, one might think about
its use for chemotoxic purposes. The numerous problems to
overcome are of course comparable to what has been describedwith the intercalators but the principal physico-chemical authenticity is given. We found that the [Re(OH2)3(CO)3]+ induces invitro irreversible conformational changes in plasmide DNA as
was previously observed for cisplatin.37, 38 The compound also
clearly reduces the viability of cells but on a higher concentration level than cisplatin. Assuming that [99mTc(OH2)3 (CO)3] +or [188Re(OH2)3 (CO)3] + would find their way into the nucleus,
an intriguing possibility of combining both radio- and chemotoxicity with one single compound would emerge. The coordination to guanine and small nucleotides includes more realisticallythe potential of direct labeling of antisense oligonucleotides asrequested for gene therapy or for the antisense strategy without
the need of conjugating a ligand to these biomolecules.
In conclusion, these three brief examples should underline
the importance of new synthons in what ever respect and as
represented by [99(m)Tc(OH2)3(CO)3]+. It should also encourage
inorganic chemists to reactivate the corresponding research withsimple compounds and to think about novel building blocks.
As shown with the example from cyclopentadienyl chemistry,
research directions with other transition metals can be initiatedand very interesting results obtained, relevant not only for technetium chemistry but also for very different fields such as catalysis. Although much remains to be done and to be proven, theexample with the intercalators stands for new directions withinradiopharmacy and exceeding the ever lasting topic of labeling
peptides with bifunctional chelators. New synthons with properties different from [99mTc(OH2)3 (CO)3] + might lead to otherapproaches which are in the worst case interesting for fundamental research. Vitamin B12 is a good example for a surprisingand unexpected finding, and likely to have a long term impact inbasic inorganic and applied radiopharmaceutical chemistry.

176 J. Nucl. Radiochem. Sci., Vol. 6, No. 3, 2005
References(1) R. Alberto, Comprehensive Coordination Chemistry II
Volume 5, (Elsevier Pergamon, 2003), p. 127.
(2) J. R. Dilworth and S. J. Parrott, Chem. Soc. Rev. 27, 43(1998).
(3) S. Jurisson, D. Berning, W. Jia, and D. S. Ma, Chem. Rev.
93, 1137 (1993).
(4) K. Schwochau, Technetium, Chemistry and Radiopharmaceutical Applications. Wiley-VCH, Weinheim, 2000).
(5) C. J. Anderson and M. J. Welch, Chem. Rev. 99, 2219
(1999).
(6) R. Alberto, R. Schibli, R. Waibel, U. Abram, and A. P.
Schubiger, Coord. Chem. Rev. 190-192, 901 (1999).
(7) R. Schibli and P. A. Schubiger, Eur. J. Nucl. Med. Mol. I.
29, 1529 (2002).
(8) P. A. Schubiger, J. Grunberg, S. M. Ametamey, M. Honer,
E. Garcia-Garayoa, P. Blauenstein, R. Waibel, I. NovakHofer, and R. Schibli, Chimia 58, 731 (2004).
(9) R. Alberto, R. Schibli, A. Egli, A.P. Schubiger, U. Abram,and T. A. Kaden, J. Am. Chem. Soc. 120, 7987 (1998).
(10) R. Alberto, Eur. J. Nuc. Med. Mol. 30, 1299 (2003).
(11) L. H. Wei, S. R. Banerjee, M. K. Levadala, J. Babich. andJ. Zubieta, Inorg. Chim. Acta 357, 1499 (2004).
(12) S. R. Banerjee, K. P. Maresca, L. Francesconi, J. Valliant,J. W. Babich, and J. Zubieta, Nucl. Med. Biol. 32, 1 (2005).
(13) S. R. Banerjee, M. K. Levadala, N. Lazarova, L. H. Wei, J.
F. Valliant, K. A. Stephenson, J. W. Babich, K. P. Maresca,
and J. Zubieta, Inorg. Chem. 41, 6417 (2002).
(14) K. A. Stephenson, S. R. Banerjee, T. Besanger, O. O.
Sogbein, M. K. Levadala, N. McFarlane, J. A. Lemon, D.
R. Boreham, K. P. Maresca, J. D. Brennan, J. W. Babich, J.
Zubieta, and J. F. Valliant, J. Am. Chem. Soc. 126, 8598
(2004).
(15) R. Garcia, A. Paulo, A. Domingos, I. Santos, K. Ortner,
and R. Alberto, J. Am. Chem. Soc. 122, 11240 (2000).
(16) R. Alberto, J. K. Pak, D. van Staveren, S. Mundwiler, andP. Benny, Biopolymers 76, 324 (2004).
(17) J. K. Pak, P. Benny, B. Spingler, K. Ortner, and R. Alberto,
Chem.-Eur. J. 9, 2053 (2003).
(18) P. Benny, J. K. Pak, B. Spingler, P. Kurz, and R. Alberto,
Abstr. Pap. Am. Chem. S. 228, U6 (2004).
(19) J. Wald, R. Alberto, K. Ortner, and L. Candreia, Angew.
Chem. Int. Ed. 40, 3062 (2001).

Alberto(20) G. Jaouen and R. H. Fish, J. Organomet. Chem. 668, 1(2003).
(21) M. J. Abrams, A. Davison, A. G. Jones, C. E. Costello, and
H. Pang, Inorg. Chem. 22, 2798 (1983).
(22) J. Bernard, K. Ortner, B. Spingler, H. J. Pietzsch, and R.
Alberto, Inorg. Chem. 42, 1014 (2003).
(23) S. Masi, S. Top, L. Boubekeur, G. Jaouen, S. Mundwiler,
B. Spingler, and R. Alberto, Eur. J. Inorg. Chem. 10, 2013(2004).
(24) K. G. Hofer, Rad. Prot. Dosim. 79, 405 (1998).
(25) T. M. Behr, M. Behe, M. Lohr, G. Sgouros, C. Angerstein,
E. Wehrmann, K. Nebendahl, and W. Becker, Eur. J. Nucl.
Med. 27, 753 (2000).
(26) G. L. Griffiths, S. V. Govindan, G. Sgouros, G. L. Ong, D.
M. Goldenberg, and M. J. Mattes, Int. J. Cancer 81, 985(1999).
(27) P. Häfliger, N. Agorastos, B. Spingler, O. Georgiev, G.
Viola, and R. Alberto, Chem. BioChem. 6, 414 (2005).
(28) S. Mundwiler, M. Kundig, K. Ortner, and R. Alberto,
Dalton T. 9, 1320 (2004).
(29) R. Waibel, I. Novak-Hofer, R. Schibli, P. Blauenstein, E.
Garcia-Garayoa, R. Schwarzbach, K. Zimmermann, R.
Pellikka, O. Gasser, A. Blanc, M. Bruhlmeier, and P. A.
Schubiger, Chimia 54, 683 (2000).
(30) R. Waibel, R. Alberto, J. Willuda, R. Finnern, R. Schibli, A.
Stichelberger, A. Egli, U. Abram, J.-P. Mach, A. Plueckthun,and P.A. Schubiger, Nat. Biotechnol. 17, 897 (1999).
(31) S. Kunze, T. Zobi, P. Kurz, B. Spingler, and R. Alberto,
Angew. Chem. Int. Edit. 43, 5025 (2004).
(32) S. Mundwiler, S. Kunze, B. Spingler, F. Zobi, and R. Alberto,
Chem. Eur. J. 11, 4089 (2005).
(33) B. Fischer, S. Benzina, V. Ganansia-Leymarie, J. Denis, J.
P. Bergerat, P. Dufour, J. Gueulette, and P. Bischoff, Can. J.
Physiol. Pharm. 82, 140 (2004).
(34) S. T. Sullivan, A. Ciccarese, F. P. Fanizzi, and L. G. Marzilli,
Inorg. Chem. 39, 836 (2000).
(35) E. R. Jamieson and S. J. Lippard, Chem. Rev. 99, 2467
(1999).
(36) F. Zobi, B. Spingler, T. Fox, and R. Alberto, Inorg. Chem.
42, 2818 (2003).
(37) F. Zobi, M. Hohl, W. Zimmermann, and R. Alberto, Inorg.
Chem. 43, 2771 (2004).
(38) F. Zobi, O. Blacque, H. W. Schmalle, B. Spingler, and R.
Alberto, Inorg. Chem. 43, 2087 (2004).