**10. The potential clinical consequences of radiation induced bystander effect**

Although direct extrapolation of data from *in vitro* experiments to *in vivo* radiotherapy is not possible, (three-dimensional structure of tissues), one could assume that the bystander effect implies a risk of post-radiation complications in healthy tissues. It is suggested that genetic instability, which takes the form of delayed reproductive death (DRD), can participate in late side effects in patients treated with radiotherapy, because of damage, increased cell loss

Intercellular Communication in Response to Radiation Induced Stress:

requires further studies.

**11. Conclusion** 

Bystander Effects in Vitro and in Vivo and Their Possible Clinical Implications 355

dose was estimated at ca. 0.6 Gy (Brenner et al., 2000), appears to have been associated with the bystander effect induced by signaling molecules in the neighborhood, and with potentially mutagenic carcinogens generated by irradiated cells, although environmental

Calculations of the equivalent whole-body dose in the case of high-energy IMRT irradiation technique (Followill et al., 1997), indicate that, in comparison with conventional radiotherapy, the risk of secondary solid cancers has increased considerably. This increase is dependent on the X-ray energy and is 1% for 6 MV, 4.5% for 18 MV and 8.4% for 25 MV compared with 0.4, 1.6 and 3%, respectively, for those same radiation energy of X-rays given in a conventional way. Furthermore, as a conclusion from this study it appears that the risk of leukemia also increases after IMTR technique. The question of secondary tumors, as a succession of radiotherapy, was investigated in several recent studies [Suit et al., 2007, Trott K-R., 2009, Tubiana M., 2009, Xu et al., 2008]. Based on epidemiological and experimental radiobiological data, Suit et al. (2007) concluded that the relationship of tumor induction risk and dose is complex and differs not only between species of animals, between individuals of the species concerned, but it may also be different for various tissues and organs. Specifically, the risk increases with dose in the 1-45 Gy range for gastric and pancreatic cancer, but is stable in the 1-60 Gy dose range for bladder cancer, and even negative for colon cancer. These phenomena are difficult to explain. They could more likely be the result of genetic instability than the effect of bystander signals at lower doses, as well as result from inhibition of signals originating from cells lethally damaged by higher doses. It seems that bystander effect can have beneficial consequences, particularly in radionuclide therapy as described above and probably in brachytherapy (Brans et al., 2006) in which tumor cells irradiated by intake or absorption of isotope energy are in the immediate vicinity of non-irradiated cells inducing in them the effect. The bystander effect can also increase damage to cancer cells during treatment with boron neutron capture therapy (BNCT) (Barth et al., 2005). As previously described, the "abscopal effect" is also an example of manifestation of the beneficial effects of irradiated cells, even at a distance from their location [Kaminski et al., 2005]. However, it is also possible that pro-survival signals, sent by lethally damaged cells, may increase the chances of survival of other, less damaged tumor cells within the field of irradiation and may pose a risk of local recurrence (Mackonis et al. (2007). The mutual communication between normal and cancer cells leading to radioprotective effect to radiation targeted cancer cells, as presented above, can also be taken into consideration. Furthermore, one can expect that the individuals exposed internally to radionuclides for routine diagnostic nuclear medical procedures might be at risk of bystander effect however, prediction whether it will be damaging or protective

Radiation induced bystander effect (RIBE) is unquestionable biological phenomenon which elicits in cells not directly irradiated but being in the neighborhood of targeted cells, or being exposed to molecular signals disclosed by irradiated ones. It has been found in variable *in vitro* and *in vivo* systems. RIBE predominate at externally applied low doses and low doserate, although many data confirm its presence at clinically used doses and radionuclide exposure. It may be either detrimental or potentially beneficial event depending on dose, dose-rate, means of irradiation, cell types and environmental conditions pointing out to its

factors, genetic background and patients' lifestyle could also significantly contribute.

and longer recovery (Hendry, 2001). Increased level of chromosome aberrations and micronuclei was detected in the head and neck cancer patients undergoing radiotherapy within a year post treatment (Gamulin et al., 2008). DRD phenomenon associated with the presence of an increased percentage of stable and unstable chromosome aberrations in lymphocytes was detected in patients irradiated because of ankylosing spondylitis even several years after radiotherapy. Furthermore, increased mortality was related to single treatment course of X-rays because of this diseases (Smith & Doll, 1982). However, other studies performed in adults many years after radiotherapy in childhood haven't shown genetic instability (Tawn et al., 2005). Neither was it shown in persons having professional contact with radiation, who have suffered internal plutonium contamination at least 10 years previous to the study (Whitehouse and Tawn, 2001). Furthermore, creation of mutator phenotype as a result of genetic instability seems to increase the probability of induction of tumors. It has been shown on an animal model that ionizing radiation induces genetic instability emerging as delayed *TP53* mutations and more frequent transformation of mammary gland epithelial cells, leading to the development of cancer (Ulrich and Ponnaiya, 1998). Compared to healthy persons, irradiated cancer patients show increased incidence of second-wave primary cancers (Boice et al., 1985, Brenner et al., 2000), although the bystander effect does not need to be the only cause of such events. It is well-known that genetic predispositions and environmental factors may have significant influence on the formation of tumors (Mohandas, 2001).

Together with modern techniques of irradiation, such as three-dimensional conformal radiation therapy (3D-CRT) or intensity-modulated radiation therapy (IMRT), the purpose of which is to reduce radiation dose delivered to healthy tissues, there is an increased risk of adverse effects resulting from a possible bystander effect, especially because in these techniques larger volumes of normal tissues are exposed to a small dose (Hall, 2006). The risk of secondary cancers is increased especially in prostate cancer (Brenner et al., 2000) and cervical cancer (Boice et al., 1985, Kleinerman et al., 1995, Chaturvedi et al., 2008, Trott, 2009). Prostate surgery and radiotherapy are methods having comparable efficacy, therefore any late consequences in the form of secondary tumors should be taken into account, especially in younger people with a perspective of long-time survival. Brenner et al. (2000) compared the incidence of second-wave primary cancers in prostate cancer patients treated with surgery only (more than 50 000) to that in patients treated by radiotherapy (more than 70 000) and observed a statistically significant, although small, increase in the risk of secondary cancers in the latter group (6%, p = 0.02). This risk was associated with dose and latent time and grew with increasing survival time, amounting to 15% for patients surviving over 5 years and to 34% for those surviving over 10 years. The emerging cancers were solid tumors, such as bladder, bowel and lung carcinomas and sarcomas, the latter within the field of irradiation. The authors did not observe leukemia cases. The risk of secondary cancers after radiotherapy of cervical cancer is comparable to that of prostate cancer. Kleinerman et al. (1995) compared the risk of secondary cancers in radiation-treated, invasive cervical cancer patients (almost 50 000) with that in a group of non-irradiated patients surviving more than 30 years and showed a 12% increase in newly-diagnosed secondary cancers, where the increase was 15% after 10 years and 26% after 20 years post radiotherapy. Cancers of colon, bladder, rectum, vagina and ovary were within the fields covered by the high-dose radiation, but there were also few cases of leukemia. However, half of secondary neoplasms accounted for lung cancer. Occurrence of cancer of the lung, the organ relatively distant from the original tumor irradiation field, in which the radiation dose was estimated at ca. 0.6 Gy (Brenner et al., 2000), appears to have been associated with the bystander effect induced by signaling molecules in the neighborhood, and with potentially mutagenic carcinogens generated by irradiated cells, although environmental factors, genetic background and patients' lifestyle could also significantly contribute.

Calculations of the equivalent whole-body dose in the case of high-energy IMRT irradiation technique (Followill et al., 1997), indicate that, in comparison with conventional radiotherapy, the risk of secondary solid cancers has increased considerably. This increase is dependent on the X-ray energy and is 1% for 6 MV, 4.5% for 18 MV and 8.4% for 25 MV compared with 0.4, 1.6 and 3%, respectively, for those same radiation energy of X-rays given in a conventional way. Furthermore, as a conclusion from this study it appears that the risk of leukemia also increases after IMTR technique. The question of secondary tumors, as a succession of radiotherapy, was investigated in several recent studies [Suit et al., 2007, Trott K-R., 2009, Tubiana M., 2009, Xu et al., 2008]. Based on epidemiological and experimental radiobiological data, Suit et al. (2007) concluded that the relationship of tumor induction risk and dose is complex and differs not only between species of animals, between individuals of the species concerned, but it may also be different for various tissues and organs. Specifically, the risk increases with dose in the 1-45 Gy range for gastric and pancreatic cancer, but is stable in the 1-60 Gy dose range for bladder cancer, and even negative for colon cancer. These phenomena are difficult to explain. They could more likely be the result of genetic instability than the effect of bystander signals at lower doses, as well as result from inhibition of signals originating from cells lethally damaged by higher doses.

It seems that bystander effect can have beneficial consequences, particularly in radionuclide therapy as described above and probably in brachytherapy (Brans et al., 2006) in which tumor cells irradiated by intake or absorption of isotope energy are in the immediate vicinity of non-irradiated cells inducing in them the effect. The bystander effect can also increase damage to cancer cells during treatment with boron neutron capture therapy (BNCT) (Barth et al., 2005). As previously described, the "abscopal effect" is also an example of manifestation of the beneficial effects of irradiated cells, even at a distance from their location [Kaminski et al., 2005]. However, it is also possible that pro-survival signals, sent by lethally damaged cells, may increase the chances of survival of other, less damaged tumor cells within the field of irradiation and may pose a risk of local recurrence (Mackonis et al. (2007). The mutual communication between normal and cancer cells leading to radioprotective effect to radiation targeted cancer cells, as presented above, can also be taken into consideration. Furthermore, one can expect that the individuals exposed internally to radionuclides for routine diagnostic nuclear medical procedures might be at risk of bystander effect however, prediction whether it will be damaging or protective requires further studies.
