“68Ga-labelling and its applications”

Vijay Kumar AM
MSc (Med), PhD

Clinical Professor
Sydney Medical School, Sydney University

Principal Radiopharmaceutical Scientist

Department of Nuclear Medicine, PET
Westmead Hospital & The Children’s Hospital at Westmead.
SYDNEY, AUSTRALIA.

Indonesian Society of NM-2018

Bandung (15th Sept 2018)
Renaissance of 68Ga-labelled peptides has given new dimension to
NeuroendocrineTumor, Prostate Ca imaging & therapy.

• availability of reliable, long-lived 68Ge/68Ga Generators

• extensive automation

• development of new macrocyclic linker based 68Ga chemistry

• huge amount of clinical data

Historic beginning of NET Imaging:

The radiopharmaceutical that was first used in patients to study human NET is
123I-Tyr3-octreotide [Krenning et al Lancet 1989; 1:242].

But, 123Ilabelled agents had several limitations:

high costs, difficult labelling procedure, bowel uptake etc.

Therefore, 111In-DTPA-D-Phe1-octreotide (111In-pentetreotide) was developed

by the Rotterdam group [published 1000 Patients study EJNM 1993;20:716].

This tracer became the first radio-receptor agent commercially available as OctreoScan,
by Mallinckrodt Medical, St. Louis, MO, USA.

It was approved in 1994 by the US Food and Drug Administration as an imaging

agent for somatostatin receptor-positive NET.
• In 1960, the first 68Ge/68Ga Radionuclide Generator was described by GLEASON
A positron cow. Int J Appl Radiat Isot 8:90-94.

• Greene WT, Tucker WD (1961) – An improved gallium-68 cow. Int J Appl Radiat Isot 12:62

• In 1964 Modified by YANO J, ANGER HO: A gallium-68 positron cow for medical use.
J Nucl Med 5:484;487.

0.005M EDTA
68Ge / Al2O3 68Ga- EDTA
(inert “68Gacomplex)

• The generator design and product may not be up to the standard, but it
served the medical needs, particularly for brain imaging at that time period
 Renaissance of Ga-68
“Cationic 68Ga” instead of inert “68Ga complexes”

• Heidelberg group described the first “concept generator” in 2004.

• 68Ge/68Ga Generator & Post-processing of elute was first described in 2004

By Zhernosekov, Filosofov, Baum….Frank Roesch et al . J Nucl Med 48:1741.

• Pioneering efforts from Obninsk, Russia, a new “Cationic 68Ga”

Generator became commercially available in the mid 2005,
instead of inert “68Ga complexes”.
Razbash et al 2005; Proc of 5th Int. conf on Isotopes, 5ICI, Brussels.
Co-ordination Chemistry

&

Bifunctional Chelators
 Co-ordination Chemistry

BFC (eg. DOTA) can be used to chelate diagnostic isotopes such as 68Ga, or therapeutic
radionuclides eg. 177Lu & 90Y and the vector on the other side.

sstr
Sst
analog
Bifunctional
68Ga,177Lu, 90Y Chelator (BFC)

BFC – DOTA, NOTA, NODAGA, TRAP etc

The BFC is the under-pinning strength behind the THERANOSTICS

Macrocyclic Bifunctional Chelators
Coordination Chemistry

6 valencies 8 valencies
Ga-DOTA Y-DOTA
Macrocyclic Bifunctional Chelators (BFC)
(Commonly used)

TRAP
Hans-Jurgen Wester & Johannes Notni , 2012
NOTA, NODAGA & TRAP

TRAP >10-20 Higher

Sp activity than
DOTA & NOTA
5000GBq/µmol
 Gallium vs. Technetium - Generators

Technetium Gallium
Ion from generator TcO4- Ga3+ (Ga-EDTA)
Solution 0.9% NaCl >0.05N HCl
Column matrix Aluminium-Oxide Variable
Half life parent Short (67h) Long (271d)
Production parent Reactor, fission Cyclotron, 69Ga(p,2n)
Breakthrough Mo-99 Ge-68,
Moly-breakthrough test Cannot be tested prior
Elution Vacuum Pump/syringe
Elution yields >80% 40-80%
Elution volume with >90% <5ml <5ml

ESRR Edinburgh 2010

 The 68Ge/68Ga generator
68Ge

e
271.0 d

68Ga

b+ 1.9
67.7 m
Ge,Zn, Fe, Ti,
Problems: 68Zn

 eluate pH: 0.1 – 1.0 M HCl 6.2 b

 generator eluate volume: 5 - 10 ml
 chemical impurities (Zn, Fe, Ti, …)
 radiochemical contamination (68Ge)
 Radio-labelling of 68Ga-DOTA-TATE
Eluent
68Ge / 68Ga Generator (5.5M HCl/5M NaCl)
0.1M HCl

Step 1 Step 2

EtOH/H2O
SCX (1:1 v/v) Saline

Step 5 Step 6
waste
Step 4

Sodium acetate buffer (pH4.5); Reactor C-18

20-50g DOTA-TATE

waste
85°C for 5min 0.22µM filter

Step 3
Radio- TLC for 68Ga-DOTA-TATE

mV

120.0

100.0 68Ga-DOTA-TATE

80.0

60.0

40.0

20.0
Free 68Ga

0.0
0:00 0:30 1:00 1:30 mm:ss
 Radio- HPLC for 68Ga-DOTA-TATE
Gradient:
Mobile Phase A: TFA (0.1%) Time %A %B
Mobile Phase B: ACN (100%)
Initial 76 24
HPLC Column: Kinetex C18, 250 x 4mm 5um
Flow rate: 1mL /min 6m 60 40
Detector: UV at 218nm
Injection Volume: 100µL
8m 60 40
Run time: 20min 15 m 76 24

Counts

5000.0

4000.0

3000.0
Retention time 7.2 min
For 68Ga-DOTA-TATE

2000.0

Retention time 3.0 min

1000.0 For free 68Ga

0.0
0:00 5:00 10:00 15:00 mm:ss
 Radio-labelling of 68Ga-PSMA
68Ge / 68Ga Generator
0.1M HCl Eluent
(5.5M HCl/5M NaCl)
Step 1
Step 2

EtOH/H2O
SCX (1:1 v/v) Saline

Step 5 Step 6
waste
Step 4

Sodium acetate buffer (pH4.5); Reactor C-18

20g HBED-PSMA

waste
85°C for 3min 0.22µM filter

Step 3
 Radio- TLC for 68Ga-PSMA

mV

120.0

100.0
68Ga-PSMA
80.0

60.0

40.0

Free 68Ga
20.0

0.0
0:00 0:30 1:00 1:30 mm:ss
 Radio- HPLC for 68Ga-PSMA
Gradient:
Mobile Phase A: TFA (0.1%) in H2O
Mobile Phase B: TFA (0.1%) in ACN Time %A %B
HPLC Column: Kinetex C18, 250 x 4mm Initial 92 8
5um
Flow rate: 1mL /min 10m 40 60
Detector: UV at 218nm 12m 40 60
Injection Volume: 100µL
Run time: 15m 12m30s 92 8

Retention time 7.2 min

For 68Ga-PSMA

Retention time 3.0 min

For free 68Ga
 Somatostatin receptor expressing tumors

Somatostatin receptor
octreotide (SMS 201-995) sstr
human SRIF-receptor subtypes Bifunctional
Chelator (BFC)

N-terminus Ligand

plasma Somatostatin receptors are over-expressed in

membrane G protein neuroendocrine tumors:

hsstr2 protein response pituitary adenoma

pancreatic islet cell tumor
C-terminus carcinoid
Neuroblastoma, pheochromocytoma
Paraganglioma
hSSTR1 > 1000 nmol / L medullary thyroid cancer
hSSTR2 0.32 nmol / L small cell lung carcinoma
hSSTR3 31.6 nmol / L
hSSTR4 > 1000 nmol / L (Reubi, 1997).
hSSTR5 7.3 nmol / L
 68Ga

Somatostatin Analogs
(vectors)
DOTA

sstr

Bifunctional
Chelator (BFC)

P. Antunes &M. Ginj & H. Zhang & B. Waser R. P. Baum & J. C. Reubi & H. Maecke Eur J Nucl Med Mol Imaging (2007) 34:982–993
P. Antunes &M. Ginj & H. Zhang & B. Waser R. P. Baum & J. C. Reubi & H. Maecke

Eur J Nucl Med Mol Imaging (2007) 34:982–993

 Comparison of 18F-FDG, 111In-OctreoScan and 68Ga-DOTATOC
[Patient: male, *1939, neuroendocrine tumor with unknown primary, multiple liver and bone metastases

18F-FDG 111In-Octreoscan 68Ga-DOTATOC

R. P. Baum & J. C. Reubi & H. Maecke- 2004

68Ga-DOTANOC: a first compound for PET imaging with
high affinity for somatostatin receptor subtypes 2 and 5

Image of the month, EJNMMI 2005

Damian Wild1, Helmut R. Maecke1, Beatrice Waser2, Jean Claude Reubi2,

Mihaela Ginj1, Helmut Rasch1, Jan Mu¨ller-Brand1, Michael Hofmann3

Eur J Nucl Med Mol Imaging (2005) :724

 68Ga-DOTA NOC

SNM Image of the Year 2008

S. Fanti et al, 2008. University of Bologna

68Ga-DOTA NOC

Multiple vertebral
paraganglioma
68Ga-DOTA-TATE

Neurofibromatosis - 2

NF2

Westmead Hospital, Sydney

 68Ga sst analogs for NET imaging:

• Neural Crest Tumors

• GEP NET Tumors
18F –FDOPA

123I– MIBG
VS
68Ga – DOTA-TATE
68Ga- Antagonists

18F - FDG
18F-DOPA (Fluorodihydroxyphenylalanine) for Imaging NET

18F-DOPA - is superior in diagnostic performance in a limited

number of precise NET types:

• Medullary Thyroid Cancer

• Catecholamine-producing tumours with low aggressiveness
• Well-differentiated carcinoid tumours of the mid-gut
• Congenital hyper-insulinism
18F-DOPA 68Ga-DOTA-NOC
18F-DOPA 68Ga-DOTA-TATE
 123I- MIBG

VS

68Ga-DOTA-TATE
 68Ga-DOTA-TATE

123I-MIBG
SRS MIBG SRS MIBG
 28

7
 Therapy

&

Therapy Monitoring
New Avenues to improve PRRT in future

• Tandem PRRT - Concurrent 177Lu/90Y PRRT

• Combined PRRT (with other treatment modalities)

– Chemotherapy (Capecitabine, Doxorubicin)

– Kinase inhibitors (Sunitinib, Sorafenib)
– Intraoperative use of probes after PRRT with 177Lu
– Intra arterial injection.

• Application of Alpha emitters (eg. 225Ac, 213Bi, and 223Ra)

• Improved peptides (eg. JR-11 Antagonists)
• Bisphosphonates
• Bisphosphonates conjugated with PSMA
Macrocyclic Bisphosphonates for

Diagnosis and Endoradiotherapy

of bone metastases

Courtesy of Frank Roesch

University of Gothenburg, Mainz, Germany
 Theranostics:
New concept

Combination of
Macrocyclic DOTA with Bisphosphonates

PET- 68Ga 177Lu Endoradio-

Diagnosis Therapy

High thermodynamic + kinetic stability with several radionuclides!

Diagnosis and therapy in the same compound!
 R. G. G. Russel, F. H. Ebetino, Osteoporosis Int. 2008; 19: 733-759.

New Generation
Bisphosphonates
adsorption affinity to HAP

ad HAP N-containing BPs develop Proton-Bridges to HAP

Zol shows to have the best angle for N-H-O interaction
 Theranostic application of [68Ga / 177Lu-DOTAZOL]

prostate cancer patient

Time point PSA (ng/mL)

baseline 1217.8
Week 1 478.0
Week 8 88.0
Week 18 62.5

distinct reduction of PSA value

after two month

single dose of
5.5 GBq [177Lu]DOTAZOL
Zoledronic acid derivatives
Patient application of [177Lu]DOTAZOL
different primary tumors
 Zoledronic acid derivatives

Prospectives so far

• Fast renal clearance and high target-to-background ratio

• Promising therapy results

Potential for α-therapy:

• Labeling and stability studies with 225Ac and 213Bi

• in vivo evaluation of [225Ac]DOTAZOL and [213Bi]DOTAZOL
Bisphosphonate PSMA conjugate

Dual Targeting

PSMA

PSMA-617

DOTA

BP
 Cumulative diagnosis and therapy of bone metastases
[68Ga]BP-DOTA-PSMA

68Ga-PSMA-11 177Lu-PSMA-617
68Ga-BPAMD 177Lu-BPAMD

[68Ga]NODAGAZOL [177Lu]DOTAZOL

Rösch, F. et al., Dalton Trans, 2011, 40, 6104 Kratochwil, C. et al., J Nucl Med, 2016, 57, 1170

Courtesy of Frank Roesch

 Summary:

• 68Ge/68Ga – Generators and automation

• Macrocyclic linkers

• Somatostatin analogues / sst Antagonists

• Bisphosphonates for theranostic applications

• Use of alpha emitters as therapy tracers

Thank you

68Ga
 Post-purification

In the case of 68Ga labelled RP:

1. Bind the eluate (Ga, Ge) to IEX (resin)

11 2. wash off impurities (solun-1)
3. harvest pure Ga-68 (solun-2)

Synthesise protein or other RP