Dr. Erik rstad, KLB room 2.11 (e.arstad@ucl.ac.

uk)

Chem 2601/2011

Molecular Imaging
Lecture 3 and 4: Introduction to Nuclear imaging and Radiochemistry

Overview (lecture 3 and 4): 1) 2) 3) 4) 5) 6) The principles of Nuclear imaging Nuclear imaging techniques Instrumentation Introduction to radioactivity Production of radionuclides Radiochemistry

Principles of Nuclear Imaging (PET and SPECT)

HO HO HO
18

O F

OH

Tracer, e.g. [18F]FDG

Nuclear Imaging techniques: 1) Positron emission tomography (PET) 2) Single Photon Emission Computed Tomography (SPECT) 3) Autoradiography

Gamma ray (511 KeV)

Positron

Gamma ray (511 KeV)

Electron

- Positron: the antimatter equivalent of an electron - Positrons are emitted from certain radioactive substances - Positrons and electrons annihilates to produce two gamma rays

- A chemical is labelled with a radioactive isotope (positron emitter) - Positrons annihilate in surrounding tissue - The resulting gamma rays are emitted from the subject

Generates 3D maps of radioactivity concentration - tomographic

Tracer labelled with gamma (single photon) emitting radionuclide

Autoradiography (imaging in vitro): Contact exposure of radioactive samples (e.g. 20 m tissue section on X-ray film) Lower resolution than fluorescence microscopy, but quantitative Requires low energy beta emission

Burton et al. (2009), TOXICOLOGICAL SCIENCES, 111(1): 131139. http://www.nationaldiagnostics.com

Instrumentation: 1) Detector principles 2) Principle of PET scanners 3) Principle of SPECT scanners

10

Detector physics and image analysis: From gamma rays to 3D image

Outgoing amplified signal

Scintillation detector: Converts gamma rays to light

Photomultiplier tube (PMT): Converts light to electricity and amplifies signal

PET camera: how does it work?

detector 1

coincidence window

detector 2

time (ns)

SPECT camera: how does it work?

No coincidence detectors are fitted with collimators to filter gamma radiation

Properties of nuclear imaging techniques:

(1) Resolution (time and space) (2) Sensitivity (3) Selectivity (4) Quantification (5) Tissue penetration (6) Invasiveness (7) Structural information (8) Functional information + +++ +++ +++ +++ +++ = non-invasive + +++

Nuclear imaging enables non-invasive quantitative imaging of biological processes in vivo

14

SPECT vs. PET

Resolution Sensitivity
(gamma detection)

SPECT 12-15mm 0.03% yes Gamma emitters half-life > 6 hours 10-13 molar

PET 4-7mm 3.0% no Positron emitters half-life < 2h 10-14 molar

dual radionuclides Radionuclides Sensitivity

(Target concentration)

Cost

$$

$$$ (5002000/scan)

Radioactivity: 1) 2) 3) 4) 5) Introduction to radioactivity Ionizing radiation Half-life and radioactive decay Specific activity Attenuation

16

What is radioactivity?

17

A brief introduction to Radioactivity:

1895: Roentgen discovered X-rays 1896: Henri Becquerel discovered rays from uranium

1897: Marie Curie named the rays radioactivity 1898: Marie and Pierre Curie discovered Polonium and Radium

2012: > 2500 radioactive nuclides are known!

Definitions: A nuclide (from nucleus) is an atomic species characterized by the specific constitution of its nucleus, i.e., by its number of protons Z and its number of neutrons N. A radionuclide is any radioactive nuclide. Isotopes are atoms from the same element (i.e. same proton number) but different number of neutrons.
Mass number = N + Z Proton (Z) number

18F-

Charge

19

Radioactivity is defined as the process in which unstable atomic nuclei spontaneously emit ionizing radiation

Types of ionizing radiation:

E = mc2
Alpha particles = He nucleus Positrons = antimatter of electrons

Beta particles = electron

Gamma rays = highly energetic photons

20

The units of radioactivity:

Historical units: Ci (curie) = 3.7 1010 disintegrations per second (= 1 g of 226Ra) mCi = 37 x 106 disintegrations per second SI units: Bq (Becquerel) = 1 disintegration per second KBq = 1 x 103 Bq MBq = 1 x 106 Bq GBq = 1 x 109 Bq Conversion factor: 1 mCi = 37 MBq

21

Ionizing radiation and energy

The energy of ionizing radiation is measured in electron volts (eV) Units: KeV = 1000 eV or MeV = 1000,000 eV For particles it is the kinetic energy (typically 100 KeV to 1 MeV) For gamma rays it is the energy of the photon NB: Gamma energies for SPECT imaging ~ 100-300 KeV For PET the gamma rays are always 511 KeV (the combined mass of an electron and a positron = 1.022 MeV)

22

Relationship between half-life, time and radioactivity

The half-life (t1/2) of a radionuclide is determined by its decay constant lambda ():

t1/2 = ln 2 /

(ln = natural logarithm, ln 2 = 0.693)

The activity of a radioactive sample at any time is:

At = A0 x e t
Where A0 is the activity at time zero and e = natural constant (2.718)

Where n equals number of whole half-lives: At = A0(1/2)n

23

Half-life and radioactive decay over time

24

Question: Carbon-11 has a half-life of 20 min. The synthesis of a tracer takes 40 min and it takes another 20 min to analyse the product before injection to a subject. The radiochemical yield is 20%. How much of the initial activity is available for injection?

Specific activity: Activity / Mass = Bq / mol Direct correlation between half-life and maximum specific activity: t1/2 = ln 2 / is the probability of radioactive decay: Low = long t1/2 High = short t1/2

The shorter the half-life the higher the maximum specific activity
26

Specific activity important terms: Samples exclusively made up of molecules containing the radioactive nuclide are carrier-free (c.f.). Samples without addition of non-radioactive carrier but containing naturally occurring isotopic dilutions are non-carrier-added (n.c.a.). Samples diluted with non-labelled molecules are carrier-added (c.a.).

27

Radiation properties: Attenuation

Range of a couple of cm in air stopped by a sheet of paper

Range of mm to cm in tissue stopped by a sheet of aluminium

long range requires thick lead for shielding

Attenuation: effect of matter

Gamma ray intensity I0 Gamma ray intensity Ix

I = I0 Ix, where I is proportional to Z3

Doubling the atomic number leads to 8 fold increase in attenuation!

http://en.wikibooks.org/wiki/Basic_Physics_of_Nuclear_Medicine/Attenuation_of_Gamma-Rays
29

Attenuation: effect of matter AND energy

Half value (in cm) for gamma rays: Biological tissues: Different stopping power of radiation e.g. lungs vs. bones

http://en.wikibooks.org/wiki/Basic_Physics_of_Nuclear_Medicine/Attenuation_of_Gamma-Rays 30

Attenuation of positrons in tissue:

NB: this defines the maximum theoretical resolution of PET!

Nuclear imaging is quantitative, because: Radioactive decay is determined by the half-life Radioactive decay is unaffected by the environment The interactions of ionizing radiation with matter follows clear physical rules and can be accounted for

32

Question: What would happen if a subject is injected with a PET tracer but scanned with SPECT camera? What would happen if a subject is injected with a SPECT tracer but scanned with PET camera?

Radiochemistry: 1) 2) 3) 4) 5) Production of radionuclides Labelling with 11C Labelling with 18F Labelling with 123I Labelling with 3H

34

Radiochemistry and production of radionuclides Examples for PET: 11C (t1/2 20.4 min) and 18F (t1/2 110 min) Example for SPECT: 123I (t1/2 13.1 h) Example for autoradiography: 3H (t1/2 12 years)

35

Production of radionuclides with a cyclotron

Reaction: Product: half-life: Decay mode:

16O 14N 14N 18O

(p,) (p,) (d,n) (p,n)

13N 11C 15O 18F 123I 99mTc 68Ga 82Rb

10 min 20 min 2 min 110 min 13.1 h 6.01 h 68 min 1.26 min

+ (positron) + (positron) + (positron) + (positron) (gamma) (gamma) + (positron) + (positron) 36

124Te(p,2n)

Generator based radionuclides:

Production of radionuclides: formation of 3H

Nuclear reactor provides neutron flux:

6Li

+n

4He

+ 3H (half-life 12 years, beta emitter)

All radionuclides for biomedical research are either produced by a cyclotron, or in nuclear reactors (directly or indirectly).
37

Radiochemistry general principles: Fast reactions High yields Reliable and reproducible reactions Few side products Simple purification Introduce the radionuclide as late in the synthesis as possible!

38

Radiochemistry important definitions:

Radiochemical yield (r.c.y): the efficiency of a labelling reaction measured as the proportion of radioactivity that has been transferred from a reagent to a product. Radiochemical yield can be decay corrected or non-decay corrected: Decay-corrected: the amount of activity in the product is corrected for the decay that has occured during the synthesis before calculation of radiochemical yield Non-decay-corrected: there is no correction for decay, so the radiochemical yield is simply the amount of radioactivity in the product divided on the amount of radioactivity in the reagent.

39

Radiochemical yield - example: Carbon-11 has a half-life of 20 min. You start the synthesis with 2 GBq of 11CO2. After 40 min you obtain 200 MBq of a tracer. The non-decay corrected radiochemical yield is: 0.2 GBq/2 GBq = 10% The decay-corrected radiochemical yield is: Decay correction: 0.2 GBq/(0.5 x 0.5) = 0.8 GBq 0.8 GBq/2 GBq = 40%

Radiochemistry: labelling with carbon-11

Advantages: Enables isotopic labelling, i.e. replacement of 12C with 11C in the molecule. The biological fate of the molecule is unchanged Very versatile labelling chemistry its carbon! Disadvantages: Short half-life (20.4 min) Only available on sites with a in-house cyclotron Alternative for biochemical applications: 14C (half-life 5730 years, beta emitter)
41

Radiochemistry: Labelling with carbon-11

Cyclotron: H2
14N
11C

Sn2 reactions: I2 ROH (alcohols)

11C 11C

H4

H3I

H3OR H3OAr H3SR H3NRH

ArOH (phenols) RSH (thiols)

11C

(p,)

11C

O2

11C

11C

O2

RMgX , SOCl2 (Grignard reagents) R11C (O)Cl RNH2 (amines) R11C (O)NHR

RNH2 (amines)

11C

RCONH2 (amides) RC(O)NH11C H 3

42

Radiochemistry: Labelling with carbon-11

[11C ]Methionine:
HS OH H2N C O

11C

H3I

H311CS OH

Base

H2N

C O

[11C ]Methionine

[11C ]Way 100635:

MgBr
11C

O2

O C

OMgBr

SOCl2

O C

Cl

RNHR
N

H3C

O N

Typical specific activities: 40-200 GBq/mol Sources of 12C: atmospheric CO2

C O
43

[11C ]WAY 100635

Radiochemistry: labelling with fluorine-18 Advantages: Near ideal half-life (110 min) and low positron energy Small size makes it suitable for labelling of small molecules Fluoride can reduce metabolism of tracers Tracers can be transported for imaging at centres without a cyclotron Disadvantages: Limited reactivity, need to protect OH and NH groups! Limited chemistry fluoride is the most electronegative of all elements
44

18F

Nucleophilic aliphatic substitutions

OAc SO2CF3 O O AcO AcO OAc

[18F]FDG:

Leaving group (triflate) Acetate (protecting group)

18F-

OAc O AcO AcO

18

HCl (aq)
OAc
HO HO

OH O

OH
18

[18F]FDG
45

[18F]Fluoride Nucleophilic aliphatic substitutions

Grierson and Shields, Nuclear Medicine & Biology 2000, Vol. 27; 143156
O S O O R

O2N

Leaving group = nosylate

18FO DMBn

Protecting group
O

MeCN, K2CO3, Kryptofix 100 C, 10 min,

N N O O

Deprotection
HO O
18

NH N O

DMTrO

Protecting group

18

38% RCY

[18F]FLT 13% RCY

46

[18F]Fluoride Nucleophilic aliphatic substitutions

OMe OMe TsO
18F-

H N O

OMe OMe H N

O S

O R O

18

F
O

H3C

[18F]Fallypride
47

Leaving group: tosylate (TsO)

O S O R

[18F]Fluoride

fluoroalkylation
H3C

Leaving group: tosylate (TsO)

OTs TsO
18F-

OTs
18

[18F]Fluoroethyltosylate - reacts similarly to 11C H3I

ONH2 HO

Base (deprotonates the phenol)

O

O18

F
O

NH2

[18F]FET (fluoroethyltyrosine)
48

Radiochemistry: labelling with iodine (123,124,125,131I) Advantages: Easy labelling chemistry Many isotopes available (autoradiography, SPECT and PET) Range of half-lives from 13 h to 60 days Disadvantages: Limited metabolic stability Large size (similar to benzene!) Limited labelling chemistry
49

Direct labelling of peptides/proteins with iodine:

O

123I-

Oxidant

123

I NHR

123I+
O

HN HO
NHR

R O

HN HO

R O

Tyrosine residue

Advantages: High yields Simple chemistry Disadvantages: Low metabolic stability No control of labelling positions (in large proteins)

50

Labelling of trialkyl tin compounds with iodine (aryl and alkene):

SnMe3 OH
123 123

I OH

H N OMe O

I , H2O2

Dilute acid OMe O

H N

[123I]IBZM

Advantages: Site specific labelling Good yields Disadvantages: Only suitable for small molecules (or indirect labelling)

51

Radiochemistry: labelling with tritium (3H or T)

Advantages: Can label almost any organic compound Ideal for isotopic labelling replacing H with tritium Widely used basic research Disadvantages: Only possible in specialised labs (custom service) Only suitable for in vitro applications

52

Labelling with 3H (also known as tritium - T):

Reduction of alkenes, alkynes and other saturated bonds with 3H gas: 2
N N T T

T2, Pd/C

Alkylation with C3H3I equivalent to 11C H3I and suitable for the same reactions! Maximum specific activity for labelling with 3H ~ 2-3 GBq/mol
53

Exercise 3: How would you label this molecule?

54

Nuclear imaging strengths and weaknesses: + Excellent sensitivity (picomolar range ) + Deep tissue penetration + Allows absolute quantification - Limited resolution (time and space) - Expensive - Involves ionizing radiation
55

Synopsis: PET and SPECT principles PET

- Positron emitters - short half-life (11C, 18F) < 2 h - Detects annihilation gamma rays - Coincidence detection for location - Higher resolution - Better for quantification - More expensive

SPECT
- Gamma emitters - Longer half-life (123I) > 6 h - Detects gammas emitted directly - Collimators for location - Can use multiple radionuclides - Lower resolution - Lower cost

Synopsis: autoradiography
Detects beta particles by X-ray film. Suitable radionuclides have long halflives 2 weeks to many years! The best results are achieved with low energy beta emitters, as energetic particles are not fully stopped by the imaging medium.

Synopsis: Radioactivity
Radioactive decay: -There main modes of decay are: alpha, beta, positron and gamma emission - Radioactivity is measured in Bq = 1 disintegration per second - Activity is defined by the half-life and the number of a radionuclide - Specific Activity is the activity per mass (in Bq / mol) Interactions of ionizing radiation with matter: -Each type of radiation interacts with matter in a unique way - Particles are rapidly stopped by matter and travel only short distances - Attenuation of gamma rays depends strongly on the gamma energy and atomic number of the absorbing matter NB: Radioactivity can readily be quantified as the activity level is unaffected by the environment and the interactions of ionizing radiation with matter can be accounted for.
57

Synopsis: Radiochemistry
Production of radionuclides: Most PET radionuclides are produced with particle bombardment in a cyclotron Some radionuclides are produced in high flux nuclear reactors Radiochemistry: Reactions should be fast, efficient and reliable Radionuclides typically introduced late in the synthesis Chemistry of carbon-11, fluorine-18, iodine-123 and tritium Carbon-11 converted to CH3I or CO2 Carbon-11 labelling typically with methylation or Grignard reactions Fluorine-18 typically introduced by Sn2 nucleophilic reactions Iodine-123 introduced by addition to tyrosine or reaction with trialkytin groups Tritium typically introduced by reduction of unsaturated bonds
58

Learning outcomes - you should understand: - The principles of radioactivity - The decay modes - The interaction of radiation with matter - The principles for imaging with PET, SPECT and autoradiography - How the decay mode, half-life and energy range of the radiation effect the suitability for imaging -The advantages and disadvantages of nuclear imaging - The chemistry of common radionuclides for PET, SPECT and autoradiography
59

Assessment you should be able to apply your knowledge of radioactivity and nuclear imaging to explain underlying principles, solve practical problems and provide rationale explanations related to: - The principles of radioactivity - The decay modes - The interaction of radiation with matter - The principles for imaging with PET, SPECT and autoradiography - How the decay mode, half-life and energy range of the radiation effect the suitability for imaging -The advantages and disadvantages of nuclear imaging - Labelling reactions with common radionuclides for PET, SPECT and autoradiography
60

Useful websites and background reading:

Radioactivity: http://en.wikipedia.org/wiki/Radioactive_decay Interaction of gamma rays with matter: http://en.wikibooks.org/wiki/Basic_Physics_of_Nuclear_Medicine/Attenuation_ of_Gamma-Rays Sensitivity of nuclear imaging: John V. Fragiono, Journal of Clinical Oncology, 26(24): 4012-4021 Autoradiography: http://www.nationaldiagnostics.com Burton et al. (2009), TOXICOLOGICAL SCIENCES, 111(1): 131139. Langstrom et al. (2007), Mol Imaging Biol, 9(4): 161-175. Charon et al. (1998), Nuclear Medicine & Biology, 25:699704.

61