Multifunctional gold nanoparticles

for image-guided radiotherapy

Gautier Laurent1, Christophe Alric2, Claire Bernhard3, Sandrine Dufort4, FrédéricBoschetti5, Valerie Bentivegna6, Nirmitha Herath7, Rana Bazzi1, François Lux2, Marie Boschetti5, Valerie Bentivegna6, Nirmitha Herath7, Rana Bazzi1, François Lux2, Marie

Dutreix7, Pascal Perriat8, Franck Denat3, Géraldine Le Duc6, Olivier Tillement2, Stéphane Roux 1

1 Institut UTINAM, Université de Franche-Comté, France.2 LPCML, Université Claude Bernard Lyon1, France.

3 ICMUB, Université de Bourgogne, France, 4 Nano-H SAS, France.

5 CheMatech SAS, France. 6 ID17 Biomedical Beamline, ESRF, France.

7 Institut Curie, France. 8 MATEIS, INSA Lyon, France

The limitations of the conventional radiotherapy

Tumor Healthy tissue

D1

X-rays

Conventional

Radiation therapy

Low difference between tumor

and healthy tissue irradiation

Not sufficiently selective

The radiosensitization: a strategy for overcoming the limitations

Healthy tissueTumor

D2<D1

X-rays

Tumor loaded with

radiosensitizing agents

Radiations are focussed in tumor

and healthy tissues are spared

A prefential targeting of the tumor is required

Why are the nanoparticles better suited for tumor targeting?

Endothelial cell

Passive

targeting

Tumor cell

Leaky vasculature of tumor

Normal cell

TumorHealthy tissue

Tight jonction

Blood vessel

Active targeting

Leaky vasculature of tumor

Stealth nanoparticleNanoparticle with

targeting moleculeSmall molecule

Molecule overexpressed

by tumor cells

Preferential accumulation of the nanoparticles in the tumor

Principle of the image-guided therapy

Isignal(t)

t

tumour

Healthy tissue

Irradiation

Physical stimulus

(X or , magnetic field, NIR light)

Nanoparticles combining

medical imaging and

remotely controlled therapy

(X or γ-ray, magnetic field, NIR light)

Tumour

Treatment limited to the tumour

Sparing of healthy tissue

Improvement of the

therapeutic activity: ☺

Intravenous Injection of

nanoparticles

In vivo imaging

Why gold nanoparticles?

Localized surface plasmonresonance

Tunable optical

Strong X-ray photons absorption (high Z)

Au ?

RadiotherapyCT imaging

Tunable opticalproperties

Facile functionalization

Additional complementary properties: control of the biodistribution, imaging multimodality

Well suited for image-guided radiotherapy

Photothermaltherapy

Synthesis and characterisation of Au@DTDTPA nanoparticles

Au

� Synthesis

DTDTPA

10

15

20

25

Core size : ~2.4 nm

7

10 g.L–1 0,1 g.L–1

M+

Au

� entrapment of cationic species in the DTDTPA shell)

� Additionnal properties owing to the

formation of metal ion chelates

AdvFunctMater 2006, JACS 2008, Radiology 2011, Nanoscale 2013

0

5

10

1 10 100

~ 2,4 nm ~ 6,6 nm

DTDTPA

Au

Labelling of implants for diabetes I by Au@DTDTPA-Gd nanoparticles

� In vitro and in vivo detection and in vivo evaluation of labelled implants

Au

Au

IRM T1 IRM T2* Imagerie X Ultrasons IRM T1 Imagerie X

Au Au

T1-MRI T2*-MRI CT US T1-MRI CT

8

AuAu

Ultrasons- β cells (production of insuline)

(�) healthy mice

(�) non-treated diabetic mice

(�) diabetic mice treated by

labelled microcapsules

Limite diabète

Au

Au

Au

AuAu

Au

- Au@DTDTPA-Gd50 nanoparticles

Microcapsules of alginate APSA+GG :

Implantation in the abdomen

(~ 6000 microcapsules)

Days

[Glu

cose

] sa

ng

(m

g.d

L–1)

US

Radiology 2011

Radiosensitizing effect of Au@DTDTPA nanoparticles

40

50

60

70

80 Unlabeled U87 cells Au@DTDTPA-Gd

50 labeled U87 cells

Au@DTDTPA labeled U87 cellsM

ean

tail

mom

ent (

a. u

.)

9

No irradiation 5 Gy 10 Gy0

10

20

30

Mea

n ta

il m

omen

t (a.

u.)

Me

an

ta

il m

om

en

t /a

.u.

Efficient radiosensitizerMTM (unlabelled cells 10 Gy) = MTM (labelled cells 5 Gy )

Radiosensitizing effect of Au@DTDTPA nanoparticles

D0

Au@DTDTPA

+

MRT

Implantation Irradiation Sacrifice

D11 D61

� Microbeam Radiation Therapy (MRT, CF 400Gy, 50 µm, ctc 200 µm)

�Intratumoral injection | Rat bearing osteosarcoma

10

MRT

Au@DTDTPA

Control

0 5 10 15 20 25 30 35 40 45 50 55 60 65

Delay after implantation (days)Delay after implantation /day

Small 2014MRT + AuDTDTPA : higher MeST

Biodistribution study of Au@DTDTPA

� in vivo MRI of healthy mice

KIDNEYS

t0-5 t0+15 t0+30 t0+45

t0: intravenous injection of Au@DTDTPA-Gd50

([Au] = 10 g.L-1)75 µmol Gd / kg

t > t

11JACS 08

BLADDER

t0-5 t0+15 t0+30 t0+45

t > t0Positive contrast

enhancement

1°) kidneys2°) bladder

Biodistribution study of Au@DTDTPA

� in vivo planar γ scintigraphy of healthy rats

Au99mTc ; 111In

Au

111In111In

Au@DTDTPA-99mTcAu@DTDTPA-99mTc Au@DTDTPA-111In

t0 + 20 min t0 + 24 h

Au@DTDTPA-99mTc

Au@DTDTPA-111In

Intravenous

injection

t0

t0 + 72 h

12

t0 + 30 min

No undesirable accumulation in the organs

renal clearance

Nanoscale 2013

100 MBq 5 - 45 MBq

Biodistribution study of Au@DTDTPA

� gamma counting and determination of gold content in the organs as a

function of time (intravenous injection to healthy animals)

15

50

60

Fra

ctio

n de

la d

ose

d'or

inje

ctée

(%

)

30 min 24 h 48 h 72 h

15

60

70

inje

ctée

(%

)

30 min 24 h 48 h 72 h

Au

ICP analysis

Au

Gamma counting

13

Cerve

auCad

avre

Cœur

Foie

Muscle Os

Peau

Poumon

s

Rate

Reins

Intes

tinSan

g

0

5

10

Fra

ctio

n de

la d

ose

d'or

inje

ctée

(%

)

Cerve

auCad

avre

Cœur

FoieMus

cle OsPea

uPou

mons

Rate

Reins

Intes

tinSan

g

0

5

10

15

Aor

gane

/Ain

ject

ée

� Strong correlation between gamma counting and ICP analysis

���� no leakage of indium-111

� Renal clearance and absence of undesirable accumulation

Renal clearance of Au@DTDTPA-M

30

40

50

60

70

80

Cum

ulat

ive

% In

ject

ed d

ose

Feces Urine

14

30min 24h 48h 72h0

10

20

30

Cum

ulat

ive

% In

ject

ed d

ose

Time after injection

Removal of the nanoparticles essentially by renal excretion

� Intravenous injection | Rat bearing brain tumor (gliosarcoma 9L)

Injection of

Au@DTDTPA-Gd50

[Au] ~ 50 mM

[Gd] ~ 5 mM

Vinj = 1,4 mL

Biodistribution study of Au@DTDTPA-Gd50

9500

10000

10500

11000

11500

12000 Tumor Healthy tissue

Sig

nal i

nten

sity

(a.

u.)

a)

t0 – 2 min

b)

c)

15

Vinj = 1,4 mL

T1-weighted MRI 7T

The most opportune moment for

inducing radiotherapy

0 5 10 15 20 25 308000

8500

9000

9500

Sig

nal i

nten

sity

(a.

u.)

Time (minutes)

b)

t0 + 5 min

Radiosensitizing effect of Au@DTDTPA nanoparticles

50

60

70

80

90

100

Sur

viva

l (%

)

� Microbeam Radiation Therapy (MRT , CF 400Gy, 50 µm, ctc 200 µm)

�Intravenous injection | Rat bearing brain tumor (gliosarcoma 9L)

16

0

10

20

30

40

50

0 50 100 150 200 250 300 350

Sur

viva

l (%

)

Days after the implantation of the tumor

NT

MRT MRT 5’pi

Small 2014

Conclusion

� high colloidal stability in biological conditions

� free circulation in blood pool without undesirable

t0+30 min

RK

U

Au@L-(Gd, In, Tc)

17High potential for image-guidedradiotherapy

pool without undesirable non-specific accumulation

� Renal clearance

� Medical imaging (CT, MRI, scintigraphy)

� Radiosensitzing effect

t0 + 30 min

AdvFunctMater 2006, JACS 2008, Radiology 2011, Nanoscale 2013, Small 2014

0

10

20

30

40

50

60

70

80

90

100

0 50 100 150 200

surv

iva

nts

(%

)

jours après l'implantation

MRT 5’ pi

MRTNT

4 min after IV injectionof of Au@DTDTPA-Gd

Acknowledgements

Gautier LaurentRana Bazzi

Olivier TillementFrançois LuxLucie SanceyCharles TruilletChristophe Alric

Géraldine Le DucElke Bräuer-KrischHerwig Requardt

Pascal Perriat

Christophe Alric

Franck Denat, Claire Bernhard

Marie DutreixNirmitha Herath

Sandrine Dufort, Cédric Louis Frédéric Boschetti

Acknowledgements

ANR P2N TheraGuIma (2011-2014)ANR RPIB MULTIMAGE (2013-2017)

BQR 2010, BQR PRES UB-UFC 2012, 2013

LTP project (2011-2014)

19

ANR RPIB MULTIMAGE (2013-2017)

EquipeX IMAPPI (2011-2019)

Défi G3N 2012