|dc.description.abstract||This thesis covers research on radiotherapy of bone cancer using bone seeking
radiopharmaceuticals i.v. applied. Such research generally has been confined to the treatment
of bone pain in people with metastatic bone cancer but on occasion also for treating cases of
osteosarcoma. It is typically used when there are multiple metastatic lesions in the skeleton,
which makes local and focal treatment impractical and systemic treatment the alternative. The
treatment is tumour specific since the radiopharmaceutical targets the area of increased
mineral turnover. This allows selective uptake and prolonged radiopharmaceutical retention in
An ideal radiopharmaceutical for the treatment of neoplastic bone disease would be a
radiolabelled compound, which would predominantly accumulate in the bone lesion with
limited access to normal bone and other organs.
Criteria governing the selection of the radionuclide are particle range, physical half-life,
gamma yield (for scintigraphic monitoring), chemistry and type of ligand. The most
commonly used radionuclides at present are phosphorus-32, strontium-89, lin-1 17m,
samarium- 153, and rhenium- 186/ 188.
Currently the available bone seeking agents which are phosphonate containing ligands lend to
localize throughout the skeleton. This thesis focuses on methylenediphosphonic acid (MDP),
1- hydroxy-ethylenediphosphonic acid (HEDP), and the novel agent polyethyleneimine
functionalised with methylene phosphonate groups (PEI-MP) from this laboratory, as well as
the octa-anion ethylenediaminetetramethylphosphonate (EDTMP). MDP and HEDP are
bisphosphonates, EDTMP a multidentate aminophosphonate. In targeted therapy EDTMP is
used in combination with 153Sm and might be binding to hydroxyapatite crystals in a different
manner than HEDP.
186Re-HEDP is used for palliation of bone pain to metastatic bone cancer. Complications
known with 153Sm-EDTMP and 186Re-HEDP treatment are myelotoxicity and in some cases a
transient increase in bone pain following treatment (flare response). In contrast to this 188Re-HEDP appears to fulfil the criteria of a good radiotherapeutic agent as
it has a stronger β-emission than 153Sm and 186Re and a shorter physical and biological half-life
which reduces myelotoxicity.
The aim of [his research is to optimise and investigate novel bisphosphonate containing
ligands for targeted radiotherapy of neoplastic bone disease using rodent and primate animal
models and scintigraphy.
The successful outcome of these studies requires that the normal primate (Papio ursinus) with
its in vivo bisphosphonate behaviour for purposes of scintigraphic monitoring, (using 99mTc-
MDP), be validated as a useful model to interpret experimental results and extrapolate to man
as was done successfully in the fracture healing experiments discussed here.
The pharmacokinetics of 153Sm –EDTMP, the well-known therapeutic radiopharmaceutical
used for palliation in painful human skeletal metastases, was determined in normal primates
by scintigraphic monitoring and compared with available human data in order once more to
establish sufficient similarity of in vivo behaviour and thus direct the meaningful continuation
of these biodynamic investigations.
A novel approach for ligand optimisation involving neoplastic tissue's abnormal blood supply
(increased permeability) and lack of lymphatics (EPR effect) was investigated, whereby
radiolabelled macromolecules accumulated selectively al the target site. The synthesis of the
macromolecule polyethyleneiminomethyl phosphonic acid (PEI-MP), and its labelling with
99mTc, as well as quality control have been described in detail. Macromolecular sizes ranged
from 3 to 300 kDa and the label 99mTc was selected to serve as tracer for the scintigraphic
biodistribution studies performed in normal adult male primates. The results allowed the
identification of a suitable size-fraction, i.e. 10-30 kDa of PEI-MP, as a bone-seeking ligand.
Molecular sizing of the polymeric PEI-MP can drastically alter its pharmacokinetic
To exploit these encouraging results, of organ sparing and speedy urine excretion a study of
bone tumour accumulation of 99mTc-PEI-MP was done in five dogs with spontaneous
occurring appendicular osteosarcomas. Mean tumour: background uptake of 4:1 was obtained
with the molecular size fraction 10-30 kDa. To fulfil a therapeutic role the most suitable ligand would have to form a stable and appropriate complex with one or more of the
therapeutic radionuclides e.g. 153Sm, 186Re and 117Sn(II).
For informed selection of such a radionuclide which would be successfully complexed with
the ligand PEI-MP for targeted delivery to osteosarcoma/metastatic bone tumours, metal ion
speciation in blood plasma was used to predict fie in vivo behaviour of the potential bone-seeking
therapeutic radiopharmaceuticals. The blood plasma model ECCLES used here,
included PEI-MP as ligand, and predicted good in vivo behaviour of 117mSn(II)-PEI-MP, but
not so when complexed with 153Sm and 166Ho. Labelling of PEI-MP with 117mSn(II) and also
with 186Re was subsequently successfully achieved.
It is known that changing the radionuclide label of a particular ligand might change the
resultant biodistribution. Therefore the biodistribution of variously molecular sized 117mSn(II)-
polyethyleneitninomethyl phosphonale complexes were investigated in the normal primate
model to establish their potential as selective therapeutic bone agents. The in vivo stability of
117mSn-PEI-MP, and reduced accumulation in the kidneys and normal bone observed from
these biodistribution studies prompted the investigation of its potential to exploit the EPR
effect due to its macromolecular nature, where bone malignancies are present. The tumour
uptake of 117mSn-PEI-MP in different types of canine osteosarcoma induced into nude mice,
was therefore studied.
The osteosarcoma model followed from subcutaneous injection of canine osteosarcoma cells
with high lung metastatic capacity (HMPOS) in some mice, and without this capacity (POS)
in another group. The former yielded faster growing non ossified tumours compared to slower
growing tumours from POS cells, with ossified tissue, both observed histologically. The high
accumulation of 117mSn-PEI-MP in the bladder wall is of great concern and a focus of
There is however no doubt that the ligand PEI-MP exhibits promising characteristics for
targeted delivery of therapeutic radionuclides to neoplastic bone lesions and essentially spares
vital organs and normal bone.
The normal primate furthermore turned out to be a useful model from which to interpret
scintigaphic results related to in vivo bisphosphonate behaviour, also for scintigaphic monitoring of the pharmacokinetics and biodistribution of bone therapeutic