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INTERACTION BETWEEN HADRONS
AND CHEMOTHERAPY AGENTS
IN HUMAN CANCER CELLS CULTURED IN VITRO
First Year Workshop 2014
Miriam Lafiandra
miriam.lafiandra@unimi.it
Supervisor: Prof. P.F. Bortignon
Co-Supervisor: Prof. D. Bettega
HADROTHERAPY WITH CHARGED PARTICLES
o Conventional radiotherapy bremsstrahlung photon beams from LINAC
o Hadrotherapy charged particles accelerated with syncrotrones or cyclotrones
Absorbed Dose:
For charged particles
Bethe - Block:
To irradiate the whole tumor thickness:
Spread Out Bragg Peak
(S.O.B.P.)
To irradiate each tumor section +
adeguate margins:
Pencil beam technology
Highly conformed dose distribution to the tumor
tumorsection
SUITABLE FOR
TUMORS
Deep seated
Close to critical organs
Resistant to conventional
therapies
Dose distribution calculated for a medulloblastoma treatment planning
CHEMORADIOTHERAPY IN RDH INFN PROJECT
Therapeutic radiations (photons/hadrons)
Chemical agents radiosensitizing action
aims: To increase tumor local control
To reduce metastasis’ formation probability
EPOTHILONE B
Potential radiosensitizing agent: Interferring with cell’s cycle, it stops cells in G2/M
(cells in this phase are very radiosensitive)
It inhibits DNA repair mechanisms in tumor cells
BIOLOGICAL SYSTEMEstablished human tumor cell-lines cultured in
vitro: characteristics do not change over time (i.e.
constant proliferative capacity)
Experiments in controlled and reproducible conditions!
Cell Lines now in study:
o Lung adenocarcinoma (A549)o Glioblastoma (U251MG)o Pediatric Medulloblastoma (DAOY)
MOST IMPORTANT BIOLOGICAL EFFECT IN RADIOTHERAPY:
LOSS OF CLONOGENIC CAPACITY IN CANCER CELLS
Survived cell after irradiation ↔ it generates a colony made up of at least 50 cells
(5-6 cell divisions)
24 h
Irradiation at ½ S.O.B.P.
(dose range = 0÷5 Gy)
Incubation37°C, 5% CO2, 90% umidity)
15 days
Clonogenic survival
vs. Radiation
dose/type +/- Epothilone B
MEASUREMENTS WITH PROTON BEAMS
Samples irradiated @ CNAO in Pavia
+ Epothilone B
Determination of Epothilone B concentration for each cell line
PRELIMINARY MEASUREMENTS
Similar to the concentration in
patient’s plasma
Equitoxic (survival ≈ 40%)for the various cell lines:
A2549 0.075 nMDAOY 0.035 nM
U251MG 0.125 nM
Clonogenic survival of A549 cells vs. Epothilone B concentration
PRELIMINARY RESULTS WITH PROTON BEAMS (1/2 SOBP 15 cm)
A549 (lung adenocarcinoma) cells (4 indipendent experiments)
Surviving curves described by
Linear Quadratic Model Protons
α= (0,60 ± 0,07) Gy-1
β= (0,04 ± 0,02) Gy-2
Protons + Epothilone B
α= (0,90 ± 0,04) Gy-1
S0= (0,36 ± 0,02)Dose Enhanchment Factor:
PRELIMINARY RESULTS WITH PROTON BEAMS (1/2 SOBP 15 cm)
Protons
α= (0,57 ± 0,05) Gy-1
β= (0,03 ± 0,02) Gy-2
Protons + Epothilone B
α= (0,88 ± 0,03) Gy-1
S0= (0,36 ± 0,03)
DAOY (medulloblastoma) cells (4 indipendent experiments)
D.E.F.2Gy = 1.5
Various analysis based on different definitions of additivity between citotoxic agents:
Steel & Peckham G. Steel, M. Peckham et al., Int. J. Radiation Oncology Biol. Phys. ,V. 5, pp
85-91, 1979 G. Steel, Int. J. Radiation Oncology Biol. Phys. , V. 5, pp 1145-1150, 1979
LamLam G. K. Y., Bull. Math. Biol. Vol. 51 pp. 293-309, 1989
Luttjeboer Luttjeboer M. et al., Int. J. Rad. Biol. Vol 86, pp. 458-466, 2010
INTERACTION BETWEEN RADIATION & EPOTHILONE B
ADDITIVITY?
SYNERGISM?ANTAGONISM?
A549 cells DAOYcells
LUTTJEBOER ANALYSIS
Synergism!
sinergism sinergis
m
WORK IN PROGRESS
comparison with photons beams:
PRELIMINARY RESULTS synergistic interaction between photon beams and Epothilone B
more measurements on proton beams with A549, DAOY cells
extension of these measurements to U251MG (glioblastoma) cells
Carbon Ions (12C6+) +/- Epothilone B :(greater biological effectiveness & better physical properties)
Citofluorimetric analysis to measure Epothilone B effects on cells cycle
Epothilone B toxicity in cells derived from normal tissues
NEXT FUTURE
THANK YOU!
SYNERGIC INTERACTION’S CONSEQUENCES
Effect of the combined treatment is greater than the sum of the effects of radiation and chemoterapy used alone
it is possible to reduce radiation dose to obtain the same effect of a standard treatment
Combining the local action of radiation and the systemic one of the chemical agent major effect inside tumor volume + reduction of metastasis’ formation probability.
Epothilone B has a major effectiveness on highly proliferating cells (such as cancer cells) selectivity
LINEAR QUADRATIC MODEL Hp: Letal DNA damages caused by radiation can be due to a
single track damages frequency directly proportional to radiation dose.
Letal damage can be even caused by the interaction of lesions due to different indipendent tracks frequency quadratically proportional to dose.
Letal lesions randomly distributed.
Epothilone B combined with radiation makes the β parameter negligeble!
STEEL & PECKHAM ANALYSIS•Isoeffect plane•For a given survival level S*:
Mode I: for every chemoterapy agent’s concentration, it is reported the radiation doses that added to the chemical agent leeds to S*.
Mode II: for every concentration, it is calcolated the dose needed to produce the same effect It is then reported the dose that added to this calculated one, leeds to S*.