LECTURE 3
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Radioprotectors/Radiosensitizers
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Radioprotectors
• Compounds which reduce the effect of
radiation on the cell or organism
– Sodium cyanide, carbon monoxide, etc. act by
reducing oxygen concentration in organs, which lowers
the effect of x-rays; they’re also toxic
• The first compounds discovered (1948)
are cysteine and cysteamine
– Experiments showed these drugs could protect animal
subjects with a dose reduction factor of 1.8 (DRF = ratio
of radiation doses with and without drug, when
measuring the same level of lethality)
3
Making a better radiation
trap
• Cysteine and cysteamine are toxic - drug dose
levels necessary for protection produce nausea
and vomiting
• Here comes the army - program initiated in 1959;
first discovery was that a phosphate cover on
the sulfhydryl group greatly reduced toxicity;
larger drug doses meant larger radiation doses
could be tolerated
• Over 4000 compounds tested - these three are
prominent:
– WR-638, or cystaphos: DRF=1.6 over 7 d, 2.1 over 30 d
– WR-2721, or amifostine: DRF=1.8 over 7 d, 2.7 over 30 d
– WR-1607, or d-CON. Marketed as rat poison.
4
How do they work?
• These and other compounds have
chemical characteristics which
allow them to protect against the
adverse effects of sparsely ionizing
x and gamma rays:
– Primarily free-radical scavenging
• These compounds become less
effective as LET increases
• Protective effect tends to parallel
the oxygen enhancement ratio
• Not the whole story, since these
compounds also have some effect
against densely ionizing neutrons
5
Radiosensitizers
• Chemical or pharmacological agents that
increase the lethal effects of radiation
• Must be present when the irradiation occurs
• Key to effectiveness in tumor control is the
presence of a differential - tumor gets sensitized
more than normal tissue
• Two categories found effective- halogenated
pyrimidines and hypoxic sensitizers
• Both work to change tumor control percentages
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Radiosensitizing Hypoxic Cells
• Hypoxic cells are resistant to x-rays; efforts have been
made to induce higher oxygen levels in tumors prior to
treatment
– hyperbaric oxygen chambers
– blood transfusions, artificial blood, patient can’t smoke
• In the 60’s, the search began for drugs that mimic
oxygen’s effect on tumors; drugs must have these traits to
be effective:
– must sensitize tumor cells without undo toxicity to
normal cells
– must be chemically stable and not readily metabolized
– must be highly soluble in water and fat and able to
diffuse large (200mm) distances
– must be effective at low doses (several Gray) for
fractionated treatments
• First satisfactory candidate was misonidazole
8
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Hypoxic Cytotoxins
• Change in philosophy: kill hypoxic cells
instead of sensitize them
• Used alone, they are chemotherapeutic
agents
• Used in conjunction with radiation, is cellkilling more effective?
• Leading compound of interest is
tirapazamine (TPZ)
10
Genetic Susceptibility
Genetic Susceptibility
Can we identify cells, individuals or subpopulations
that are genetically susceptible to radiation?
Some individuals are more
sensitive than others to a
variety of things
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Dust
Animal hair
Chemicals
Sun
Drugs, medicines
Foods
Radiation
We know that radiation is one
of the things that has a wide
range of sensitivities
Resistant
Individuals
Sensitive
Individuals
Radiation Dose
Radiosensitive cells have been
developed. After the same amount of
radiation, they have more changes
than normal cells.
Normal Cells
Sensitive Cells
Sensitive and Insensitive Mice
Hybrid Mouse
Models
Some strains of mice such as BALBc are
more sensitive to radiation than others.
For example, C57BL/6 mice are
particularly resistance to radiation-induced
mammary cancer.
Survivors of radiation exposure have
demonstrated that some people are less sensitive
to radiation exposure than others.
LD50 for radiation for humans is about 300,000 mrem.
This means that at this high dose, half of all people will
die- but half of all people will still survive.
A-BOMB
Some survivors received more than 300,000
mrem, 60 years after the exposure, 40% of the
population of A-bomb survivors are still alive.
CHORNOBYL
One survivor in control room received 550,000 mrem
Why are these people apparently unaffected
by the effects of radiation?
Genetic susceptibility can be passed on
from one generation to the next,
therefore it probably involves genes.
• Strains of mice have been developed that are more
sensitive to radiation than others.
• Cell lines have been developed that are more
sensitive to radiation than others.
• People with some genetic diseases, such as Ataxia,
are radiation sensitive.
Multiple genes contribute to
radiosensitivity
• Different genes respond to high radiation and low
radiation. The types of genes vary.
• Most biological systems have back ups or require
homologous chromosomes, so that one mutation or
irregularity does not automatically cause a problem.
• Most sensitivity to radiation involves disruptions of
multiple genes.
Genes which may effect
Genetic Susceptibility
• Radiation-induced genes
– Some genes are activated or deactivated by radiation- these genes
may make people more sensitive or more resistant to radiation
damage.
• Stress response genes
– If these genes cannot deal appropriately with oxidative stress caused by
radiation, the function of the cell can be disrupted.
• DNA repair genes
–
Most radiation damage to DNA is repaired. If DNA repair genes are defective
then cells cannot fix even minor damage caused by radiation.
• Apoptosis genes
–
Genes which trigger the normal death of cells may malfunction, resulting in
inappropriate death or survival of altered cells.
Researchers have developed
methods to identify radiation
sensitive and resistant individuals
• Changes in gene expression are being used to predict
sensitivity in individuals.
• It has been found that people with increased
radiation-induced aberrations at the G2 stage of the
cell cycle are more sensitive to radiation therapy.
• Dose response for cells taken from patients can help
predict their radiation sensitivity.
The impact of genetic
susceptibility
• Identification of sensitive subpopulations may suggest
an increased risk at low doses for that unique
subpopulation.
• It might then be possible to control environmental
exposure to these sensitive subpopulations.
• Resistant individuals would have lower than
average risk.
Summary
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Radiation does not effect individuals to the same
degree.
•
Some people may be radiosensitive, while others
may be more resistant to the effects of radiation.
•
Scientists are trying to find better ways to
determine if someone is particularly sensitive to
radiation.
•
Understanding genetic susceptibility will help
predict and control risk in clinical and occupational
settings.
Retrieved from: http://www.pagepulp.com/936/ode-to-a-nightinga
Retrieved from:
Ode to a Nightingale
Thou wast not born for death, immortal
Bird !
No hungry generations tread thee down
;
The voice I hear this passing night was
heard
In ancient days by emperor and clown.
Breast Pathology
Breast Anatomy
Retrieved from: http://www.stfranciscare.org/saintfrancisdoctors/cancercenter/nci/popUpDefinitions.aspx?id=CDR304766.xml
Cancer Progression
Retrieved from: http://www.cancer.org/cancer/breastcancer/detailedguide/breast-cancer-what-is-breast-cancer
understanding of cancer genetics. One of these models is the first genetic
model for colorectal tumorigenesis (Fearon and Vogelstein 1990, Figure 1-1).
Multistage carcinogenesis
Chromosome: 5q
12p ; 18q
17p
Alteration:
Mutation or loss
Mutation ;Loss
Loss
APC
KRAS ; DCC
TP53
Gene:
Normal
Epithelium
Early
Adenoma
Late
Adenoma
Other alterations
Carcinoma
Metastasis
Figure 1-1. A genetic model for Colorectal Tumorigenesis. Tumorigenesis
proceeds through a series of genetic alterations involving oncogenes (KRAS)
and tumor suppressor genes (APC, TP53; particularly those on chromosomes
5q, 17p, and 18q). Adapted from Fearon and Vogelstein (1990).
The principles of Fearon and Vogelstein’s genetic model for colorectal
tumorigenesis are as follows: first, colorectal tumors appear to arise as a
Very complicated biology but
simplistic models
88.7%
• 47 out of 53 medical research papers
on the subject of cancer were
irreproducible
Sources: Nature 483 (7391) 531-533
PLOS one 8(5) e63221
•Cellular Response
•Micro-environment
•Therapeutic Advantage
•Consistency
•Individual
•Whole Organism
•Population
•Ecosystem
•Transgenerational
What is system biology ?
As Denis Noble pointed out, although there is as yet no clear definition
of the term systems biology (Noble 2005)…
• a consensus is emerging that it represents an approach to biological
research that attempts to provide a quantitative theoretical description of
the manner in which biological and physiological function emerges from
interactions between system components across multiple spatial and
temporal scales.
• It is necessarily both interdisciplinary and fully grounded within
experimental data, and as such can be seen to reflect a pioneering spirit.
The Law of Bergonie and Tribondeau
(1906)
• “X-rays are more effective on cells
which have a greater reproductive
activity;
• the effectiveness is greater on those
cells which have a long lineage;
• on those cells the morphology and the
function of which are least definitively
fixed.”
– [1], [2]
Radiosensitivity Dictated by the Kinetics
of the Mitotic Cycle
– Sinclair’s results on CHO cells:
• Cellular response to ionizing radiation is
dependent upon cell cycle phase in
which it exists during irradiation [4]
– Short et al.’s results in human cells
[5]:
• In agreement with the findings of Sinclair
– Clowes’ results:
• The length of M, G2, S phases are fairly
similar among different cells; the length
of G1 dictates the length of the total cell
cycle time [6]
◦ Therefore…
 Quickly proliferating cells can be presumed to be more
radiosensitive due to their frequency within the radiosensitive G2
phase as compared to slower-proliferating cells
 Supports the Law of Bergonie and Tribondeau
Lymphocytes – the exception to the rule
Lymphocyte
– particularly radiosensitive white blood cell arrested in G0
– nuclear material configuration of G0 cells are representative of G1 phase
– Do not transition through G2 and M phases therefore should not be
particularly susceptible to radiation; the contrary is true
Cell Survival curves demonstrating the sensitivity of various
human cells to low LET ionizing radiation
10
1
G0 lymphocytes
Surviving Fraction
•
0,1
HF19 Fibroblasts
Human epithelial cells
0,01
HCT8 Colon Adenocarcinoma cells
0,001
HT29 Colon Adenocarcinoma Cells
0,0001
0
2
4
Dose (Gy)
6
8
Survival Curve data from [7, 8, 9]
Tumour Cells
• 7 human tumour cell lines
– Looked for a relationship between the radiosensitivity of each
cancer cell line and its corresponding doubling time [10]
Cell Line
Type of Cell
HT29
HeLa
MCF7
MeWo
SW48
HCC1937
CAPAN-1
Colorectal Carcinoma
Cervical Carcinoma
Breast Carcinoma
Melanoma
Colorectal carcinoma
Ductal Carcinoma
Ovarian Carcinoma
Least Radiosensitive
Most Radiosensitive
Surviving
Fraction at 2 Gy
(%) [23]
74 ± 2
72 ± 3
68 ± 2
22 ± 2
22 ± 6
13 ± 3
9±3
Doubling Time
(h)
Doubling Time
references
23 h
19 h - 24 h
39-54 hrs
23.8 ±2.5 h
35 – 38 hrs
61 h
41 h – 97 h
[91]
[66, 69]
[10, 106]
[58]
[82, 92]
[93, 108]
[28, 29]
Law of Bergonie & Tribondeau
CAPAN-1
HCC1937
MCF7
SW48
MeWo
HT29
HeLa
Chavaudra et al. [10]
HT29
HeLa
MCF7
MeWo
SW48
HCC1937
CAPAN-1
A Different Perspective
• Tissue sensitivity as opposed to individual cell sensitivity
• Casarett’s Classification of Tissue Radiosensitivity [24]
– “radiosensitive” tissues
• Parenchymal compartment: VIM and DIM cells
• Direct damage to parenchymal cells by radiation
– “radioresistant” tissues
• Parenchymal compartment: RPM or FPM cells
• Radiation damage accumulated by more sensitive supporting
structures (MCT cells)
Cell Radiosensitivity
Classification
Relative
Radiosensitivity
Rate of Division
Differentiation
Life-span
Vegetative Intermitotic cells
(VIM)
High
Divide regularly
Poorly differentiated
Short-lived
Differentitating Intermitotic
cells (DIM)
Fairly high
Divide regularly
More differentiated than
VIM
Short-lived
Multipotential connective
tissue cells (MCT)
Intermediate
Divide irregularly,
sporadically in response to
stimuli
Variable
Variable
Reverting Postmitotic cells
(RPM)
Fairly low
Do not undergo division
regularly, only in response
to stimuli
Some are highly specialized
while some are not
Long-lived
Fixed Postmitotic cells (FPM)
Low
Do not divide, or have lost
the ability to divide
Highly differentiated
Long-lived
Conclusions
• Overwhelming evidence of non-compliance with
the Law of Bergonie and Tribondeau
– Tumour cells represent a large population of defiant
cells despite Bergonie & Tribondeau’s initial
assumptions
• “From this law, it is easy to understand that roentgen
radiation destroys tumours without destroying healthy
tissue”
• - Bergonie & Tribondeau, 1906 [1, 2]
• Law of Bergonie & Tribondeau is inadequate in
defining intrinsic radiosensitivity among the entire
cell population
– Its use will not benefit in the clinical setting when the
goal is to use our knowledge of radiosensitivity to
eradicate tumour cells
Neutron data
No neutron bystander effect in vitro or vivo
Z. Liu, C Mothersill, F McNeill, SH Byun, C Seymour W Prestwich, A dose threshold for a medium transfer bystander effect in a skin cell line,
Radiat. Res.(2006) 166, 19-23
Z. Liu, S.H. Byun, W.V. Prestwich, C.E. Mothersill, C.B. Seymour, F.E. McNeill,
Fluence and dose measurements for an accelerator neutron beam,
Nucl. Instr. Meth. B., Vol. 263, (2007), pp. 326-328.
C. Wang, SH Byun, R Smith F McNeill, W Prestwich, C Mothersill and CB Seymour, Neutrons do not produce a bystander effect in zebrafish
irradiated in vivo
Radiation research submitted
Know your target!
• Radiosensitisers failed to improve
therapy outcome in humans because it
was assumed the sole target was DNA
– Focused on hypoxic cells (less free
radicals)
– Inhibiting DNA synthesis (less tumour
growth)
• System biology and system adaptability
were ignored
Wislawa Szymborska
Poetry
- 16 Published Poetry Collections
Wislawa Szymborska
(1923-2012) Poet
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Jagiellonian University (Poland)
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[1]
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latego zyjemy (1952)
Pytania zadawane sobie (1954)
Wolanie do Yeti (1957)
Sól (1962)
Wiersze wybrane (1964)
Poezje wybrane (1967)
Sto pociech (1967), Poezje (1970)
Wszelki wypadek (1972)
Wybór wierszy(1973)
Tarsjusz i inne wiersze (1976)
Wielka liczba (1976)
Poezje wybrane II (1983)
Ludzie na moscie (1986).
Koniec i poczatek (1993, 1996)
Widok z ziarnkiem piasku. 102 wiersze (1996).
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Notable Poems:
Literature and sociology
Worked as an editor and columnist
(Krakow literary publication)
Recipient of Nobel Prize in
Literature (1996)
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Utopia
On Death, without exaggeration (O smierci
bez przesady)
Possibilities (Mozliwosci)
[1] Retrieved from: http://mlodzi.wroclaw.pl/sites/mlodzi.wroclaw.pl/files/users/22/850/6a50bf39317308d829e95416283b3a99
Eksperyment
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Eksperyment
Jako dodatek przed wlasciwym filmem,
w ktorym aktorzy robili, co mogli,
zeby mnie wzruszyc, a nawet rozsmieszyc,
wyswietlano ciekawy eksperyment
z glowa.
Glowa
przed chwila jeszcze nalezala do —
a teraz byla odcieta,
kazdy mogl widziec, ze nie ma tulowia.
Z karku zwisaly rurki aparatu,
dzieki ktoremu krew krazyla nadal.
Glowa
dobrze sie miala.
Bez oznak bolu czy chocby zdziwienia
wodzila wzrokiem za ruchem latarki.
Strzygla uszami, kiedy rozlegal sie
dzwonek.
Wilgotnym nosem umiala rozroznic
zapach sloniny od bezwonnego niebytu
i oblizujac sie z wyraznym smakiem
toczyla sline na czesc fizjologii.
Wierna psia glowa.
poczciwa psia glowa,
gdy ja glaskano, przymruzala slepia
z wiarą, ze nadal jest czescia calosci,
ktora ugina pod pieszczota grzbiet
i wymachune ogonem.
Pomyslalam o szczesciu i poczulam strach.
Bo gdyby tylko o to w zyciu szlo,
glowa
Experiment
As a supplement to the competent film,
in which the actors did what they could
to make me shrug, and even make you laugh,
displays an interesting experiment
with his head.
Head
before the hour had belonged to and now was cut off,
anyone could see that there is no trunk.
tubes dangling from his neck camera,
thanks to which the blood still circulated.
Head
had good August.
No signs of pain or even surprise
she continued to scan the movement of the torch.
pricked his ears when he heard the bell.
Damp himself he could distinguish
the smell of lard from odorless oblivion
, and licking his lips with a pronounced taste
waged spit on the part of physiology.
Faithful dog's head.
honest dog's head,
when I was stroking, przymruzala blind
with faith that is still part of the whole,
Which one yields the caress back
and wymachune tail.
I just thought about happiness and I felt fear.
because if only that were going in your life,
head
to be happy.
- Wislawa Szymborska