Purpose: To review the available clinical dose response data for extramedullary plasmacytomas (EMP) and solitary plasmacytomas of the bones (SPB), including standard 12 Gy TBI treatments for multiple myeloma (MM), to compute the expected dose response for plasma cell neoplasms and evaluate differences between EMP and SPB dose response. Method and Materials: Articles from 27 published studies on plasmacytomas were analyzed. Local control (LC) was used as the end point. Clinical data are often reported as LC for the median of a dose range — only data from ranges of width ⩽10 Gy were used. The maximum likelihood method (ML) was used to estimate the parameters of a tumour control probability (TCP) model based on Poisson statistics, and approximate likelihood confidence regions (CR) were determined. A Monte Carlo experiment (MC) assessed the parameters' uncertainty due to the 10 Gy dose interval. A statistical test based on the ability of the MC distributions of the parameters to discriminate between different kinds of tumors was performed. Results: Radiation therapy was used as the sole treatment in more than 70% of the patients and in 8 of the 12 studies selected. Parameters characterizing TCP and 95% confidence intervals from MC are reported, along with graphical representations of the dose response, and 2D MC histograms and the CRs on the parameter space. Conclusion: An extensive review of plasmacytoma clinical data was performed. Although the data suffer from a lack of low dose data and are mostly reported within a dose range, this approach is a preliminary assessment of dose response relationship for plasma cell neoplasms. The parameters of the TCP model were determined. Significant difference was seen between EMP and SPB dose response. The models could be used to interpolate clinical data and estimate TCP when assessing new therapies and comparing different treatment planning approaches.

Purpose: To examine junctioning of longitudinally adjacent PTVs treated with helical tomotherapy (HT). Method and Materials: Cylindrical PTVs were defined in an elliptic cylindrical homogeneous phantom. Dose distributions (95% PTV to receive 2 Gy) created using 2.5 and 5.0 cm long HT fields were calculated and verified dosimetrically. Cranial — Caudal (CC) dose profiles were summed to study the junctioning of PTVs to create a single contiguous PTV. Junctioning adjacent PTVs with different inter-PTV spacing, created by equal or different field sizes was studied for dose homogeneity. The use of dose stepped PTVs near the junction region was also examined. Here the SUP end of the INF PTV or the INF end of the SUP PTV was divided into smaller subPTVs of decreasing prescription dose. The resulting dose distributions were summed as a function of inter-PTV spacing. Simulated dose profiles were verified by film dosimetry. Results: The most homogenous dose resulted when adjacent PTVs had the same CC dose profile (field size). Independent of the Inter-PTV spacing, PTVs of different CC dose profiles could not produce homogeneous doses. Minimizing the volume dose excursion from prescription resulted in cold spots (−26%) and hot spots (+29%) with 8% of the PTV receiving < 95% of prescription. Dividing each PTV into four multiple contiguous subPTVs, with constantly decreasing prescribed dose (2, 1.5, 1.0, 0.5Gy) allowed PTV matching with dose homogeneity similar to junctioning PTVs of equal CC slope. 95% of the PTV received at least 101% of the prescribed dose, with dose excursions of −19% to +13% from prescription, (1% of the PTV received less than 95% of prescribed dose). Conclusion: Junctioning adjacent PTVs is possible, but PTVs created by different field widths present a challenge. Homogeneity is improved by breaking PTVs into multiple contiguous subPTVs modified to feather (broaden) the effective junctioning region.

The purpose of this study was to establish whether a pulsed dose rate (PDR) treatment of 1.5 Gy given every 3 h in combination with 41 degrees C mild hyperthermia or a continuous low dose rate (LDR) treatment with mild hyperthermia could radiosensitize two isogenic human breast carcinoma cell lines in comparison to pulsed dose rate or low dose rate irradiation alone. The radiation resistant cell line was derived from the parental cell line and was transfected to over-express DNA polymerase beta. The end-points assessed were the survival of the cells using the clonogenic assay, the amount of residual DSB(s) using the comet assay and gene expression of polymerase beta using RT-PCR. Results showed that the PDR and LDR treatments combined with mild hyperthermia caused significant radiosensitization when compared to PDR and LDR irradiation alone in terms of the clonogenic and comet assays with both cell lines. RT-PCR results showed that polymerase beta levels of expression were not elevated in response to these treatments, implying that this polymerase may not be involved in sub-lethal damage repair or thermal radiosensitization. These results suggest a potential clinical advantage when combining LDR or PDR with hyperthermia, since they indicate that hyperthermia is an effective radiosensitizer.

Cell lines mutant in specific DNA repair pathways were used to determine if these pathways are involved in adaptive responses. For these studies, the effect of deficiencies in homologous recombination repair (HR) were studied in the parental AA8 and mutant irsISF cell line pair and for deficiencies in the nonhomologous endjoining (NHEJ) pathway in the mouse MEF parental and Ku80 mutant cell line pair. The results showed that the XRCC3 mutation in the HR-deficient mutant inhibited adaptive responses to low doses of cisplatin and radiation. The parental lines showed transient adaptive responses to both low-dose cisplatin and radiation treatment. For the mouse MEF and the Ku80 cells, no adaptive responses were observed in either cell line. However, there was an initial transient sensitization response followed by partial recovery. Thus, it appears that the HR repair system may be involved in the adaptive response to cisplatin and radiation. For the NHEJ repair system the question could not be answered since no adaptive responses were evident in the parental line.

Hyperthermia has been shown in many studies to be a strong sensitizer for cisplatin treatment and this sensitization may be in part due to the inhibition of DNA repair processes. We have set out to test this in cells with specific gene knockouts for known repair processes. The chicken DT40 cell system was used with a parental line (DT40) and knockouts of homologous recombination (HR) repair DT40Rad54, nonhomologous recombination endjoining (NHEJ) repair (DT40Ku70) and a double knockout mutant DT40Ku70Rad54. The results show that thermal cisplatin sensitization was achieved in all cell lines when hyperthermia at 45 degrees C for 1.5h was given before cisplatin treatment and 42 degrees C hyperthermia was given concurrently with cisplatin treatment. The data show that inhibition of the HR repair system did not significantly affect sensitization, while inhibition of NHEJ reduced thermal sensitization at low cisplatin doses and short treatments and for concurrent treatments. These data indicate that there may be a partial involvement of NHEJ in thermal cisplatin sensitization under specific treatment conditions.

The aim was to investigate the effect of combined treatment of cisplatin with acute or pulsed radiation in human ovarian carcinoma cells sensitive and resistant to cisplatin. Human ovarian cancer cell parental line A2780s and a derivative cisplatin-resistant line A2780cp were given cisplatin treatment before a single acute dose irradiation or concurrently during a pulsed-dose irradiation sequence. Cells were irradiated in the confluent state, trypsinized and plated after treatment. When the combined treatment for cisplatin was given before acute irradiation, the results showed additive to superadditive effects for both cell lines. However, the superadditivity was only significant in the sensitive cell line. For the concomitant treatment of cisplatin during pulsed-dose irradiation the results were additive, except for the highest cisplatin dose in the A2780cp line where subadditivity was observed. The results indicate that the combined treatments could be clinically useful even though the results are mostly not superadditive. However, high-dose cisplatin (3 mu g ml(-1)) caused a subadditive effect in the resistant cell lines for pulsed irradiation. Thus, high-dose cisplatin to overcome resistance is not effective. Cisplatin with both acute and pulsed irradiation showed additive effects indicating no advantage of using cisplatin in pulsed irradiation where sublethal damage repair may be greater.

The effect of protracted mild hyperthermia treatment at 40 and 41 degrees C given, concurrently with cisplatin, was evaluated in human normal AG1522 and human mutant XPA cells. While mild hyperthermia itself for up to 6 hours showed little to no toxic effects, it did result in significant sensitization of response to cisplatin treatment. Sensitization for the normal and mutant cell line was comparable, indicating that nucleotide excision repair (NER) probably does not have a role in this process. For the 41 degrees C heating, thermotolerance developed and heating times greater than 4 hours resulted in protective effects from cisplatin cytotoxicity. This was not observed for heating at 40 degrees C for up to 6 hours.

The responses of cells with mutated DNA repair pathways were compared for cisplatin, radiation and combination treatments. The knockout of the nonhomologous endjoining (NHEJ) pathway resulted in increased radiation sensitivity, but no change in cisplatin response in the mouse cells and increased radiosensitivity but decreased cisplatin sensitivity in chicken cells. The mutation of the homologous recombination repair (HR) pathway through XRCC3 in CHO cells resulted in increased radiation and cisplatin sensitivity and to a lesser extent for the Rad54 knockout in the DT40 chicken cells. The combination treatments of cisplatin and radiation showed that inhibition of the HR repair pathway resulted in super additive effects while the inhibition of the NHEJ pathway in DT40 had no effect. In mouse cells the knockout of the NHEJ pathway resulted in reduced super additivity compared to the parental cell lines. These data show that radiation, cisplatin and combination treatment damage is affected differently by the various DNA repair pathways, which could have a range of effects on combination treatments in tumour cells expressing different levels of DNA repair in the various repair pathways.

Earlier data showed that cultured cells are more hyperthermia sensitive in S phase of the cell cycle. In part this sensitivity may be related to the disruption of DNA repair/processing activity in S phase. It is well known that many components of DNA repair systems are in fact involved in DNA replication/processing. For this study we set out to evaluate a wide range of DNA repair mutants to determine their thermal responses compared to the wild-type cells. Mutants of nucleotide excision repair (NER), homologous recombination repair (HR) and nonhomologous endjoining (NHEJ) were studied. In all cases the mutants were more thermally sensitive than the wild-type counterparts. In addition, studies on thermal tolerance (TT) showed that mutants did not have a smaller TT response than the wild type thus ruling out TT as a cause in the increased sensitivity. Cell cycle analysis showed no significant difference amongst mutant and wild-type cell line pairs and thus effects of differing cell cycle distribution was also ruled out. It is speculated that it may be the involvement of the mutated pathways in DNA replication/progressing that make mutated cells more sensitive than the wild types.

The effect of mild hyperthermia on cisplatin sensitization was examined in two cell line pairs, CHO parental AA8 and irsISF, an XRCC3 mutant (deficient in homologous recombination repair), and mouse parental MEF and knockout Ku80 mutants (deficient in non-homologous endjoining repair). The results showed that mild hyperthermia 40, 41 and 42 degrees C given concurrently with cisplatin treatment caused significant sensitization. The degree of sensitization was comparable for the parental and mutant lines, indicating that these repair pathways were likely not involved in cisplatin thermal sensitization. The shorter concurrent treatments cause a larger sensitization than the longer treatments. The reasons for this are not clear, but thermotolerance may be a factor.

Thermal radiosensitization has been shown to cause inhibition of repair of sublethal and potentially lethal damage and DNA DSBs. In this study we assessed thermal radiosensitization in mutants deficient in homologous recombinational (HR) repair and nonhomologous end joining repair (NHEJ). Using cells of the mouse wild-type embryo fibroblast cell line MEF and its Ku80(-/-) derivative that is deficient in NHEJ, we showed that thermal radiosensitization is the same in both cell lines. Further studies with cells of the wild-type CHO-AA8 cell line and its derivative IRSISF which is deficient in HR, also showed comparable thermal radiosensitization in both cell lines. Further experiments using cells of chicken DT40 cell lines also showed comparable thermal radiosensitization between the wild-type HR mutant Rad54, the NHEJ mutant Ku70, and the double mutant Rad 54-Ku70. These results indicate that the HR and NHEJ pathways may not be targets for thermal radiosensitization. (C) 2004 by Radiation Research Society

Three pairs of human tumour cell lines, with one line of each pair resistant to cisplatin, were used to compare the effects of cisplatin and ZD0473 on cellular toxicity and radiosensitization. Whilst all three cell line pairs had one line that was resistant to cisplatin, for ZD0473 the lung tumour HTB56 cp and cervical carcinoma ME180 cell lines did not express resistance to their HTB56 and SHA counterparts, respectively. Only the ovarian carcinoma line A2780 cp showed resistance to ZD0473 compared to its counterpart A2780 s . For radiosensitization both cisplatin and ZD0473 show additive and subadditive effects in the ovarian carcinoma lines, and additive and superadditive effects in the cervical carcinoma and lung tumour cell lines. In fact in the lung tumour cell lines ZD0473 appeared to be a more effective radiosensitizer than cisplatin.

Thermal radiosensitization was tested in a pair of mouse cells (MB+ wild-type and MB-, DNA polymerase beta knockout cells) and in human breast carcinoma cells (MCF7 wild-type and C716 transfected to give elevated DNA polymerase beta expression). Results showed that neither reducing DNA polymerase beta (involved in base excision repair) nor increasing it had any significant effect on thermal radiosensitization. The data indicated that polymerase beta was not involved in thermal radiosensitization, and since hyperthermia is known as a radiation damage repair inhibitor, other repair pathways might be involved and need to be explored.

PURPOSE:Human gliomas are known to be radioresistant and the aim was to determine if this resistance in part could be due to an adaptive response. MATERIALS AND METHODS:Human U-87MG glioma cells were used. Three different radiation regimens that could be related to clinical treatments were tested for their ability to cause an adaptive response. Cell survival and DNA double-strand breakage were the measured endpoints. RESULTS:All three regimens caused an adaptive response in terms of cell survival when given priming doses of radiation. The DNA double-strand break endpoint also showed fewer breaks when the adaptive response occurred. CONCLUSIONS:Using irradiation regimens that closely resembled clinical applications, in vitro data are presented that show an adaptive response in human glioma cells. This effect in part could be responsible for the radioresistance of human gliomas.

Background and purpose: A small proportion of patients undergoing radiotherapy display heightened normal tissue reactions, we have set out to determine whether this sensitivity is genetic in nature and can be assessed using an in vitro skin fibroblast assay in order to predict and avoid excessive normal tissue complications.Patients and methods: In this study we compared fife arteriovenous malformation (AVM) patients who were treated with radiotherapy and showed severe normal tissue reactions (necrosis) to two AVM patients who showed normal reactions. Fibroblasts taken from patients were cultured in vitro and irradiated.Results: The results showed that the fibroblasts from the sensitive patients were also more radiosensitive in vitro than the cells from the patients. nor-mall responding.Conclusions: The results suggest underlying genetic radiosensitivity and that such an assay may be used for prediction of severe radiosensitivity in AVM patients. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.

Purpose : To investigate the extent of non-homologous end-joining (NHEJ) in the mechanism of cisplatin radiosensitization. Materials and methods : Ku80-deficient cells are deficient in the non-homologous DNA double-strand break repair process, while the wild-type MEF cells maintain full mammalian cell repair capabilities. Both cell lines were exposed to clinically applicable doses of cisplatin (1, 3 and 6 μ g ml -1) for 1 h immediately before exposure to 250 kV X-rays. Radiation responses were plotted for each cell line and for all doses of cisplatin to observe relative levels of radiosensitization. Split-dose experiments were also performed on each cell line to measure levels of sublethal damage repair. Results : Radiosensitization was observed in the wild-type cells but not in the Ku80 cells when treated with a combination of 1 μ g ml -1 cisplatin followed by X-rays, implying that the NHEJ pathway may play a large role in cisplatin radiosensitization. Concurrent administration of this cisplatin dose with radiation produced similar levels of radiosensitization. Conversely, 3 and 6 μ g ml -1 cisplatin applied immediately before radiation revealed an increasing resistance to radiation in both cell lines -- possibly due to resistant subpopulations of cells remaining after subsequent lethal doses of cisplatin. Further experiments revealed that the high concentrations of cisplatin did not alter cell cycle distribution. Finally, split-dose experiments revealed that the NHEJ pathway also plays a significant role in sublethal damage repair. Conclusions : The study reveals that clinically applicable doses of cisplatin treatment results in the radiosensitization of mammalian cells due to the inhibition of the operation of NHEJ.

The role of polymerase beta in response to radiation, cisplatin and hyperthermia was examined in a pair of mouse cell lines, comprising a normal parental line and a derivative with polymerase beta knockout. Cell survival was assessed using the colony survival assay. For irradiation, there was no difference in response between the two cell lines. Treatment with cisplatin for 1 h showed a large increase in resistance in the mutant cell line. The results with hyperthermia were more complex. The mutant was more resistant to 45degreesC heating, but was slightly more heat sensitive than the wild type at 41degreesC. Thus, in summary, while the knockout of polymerase beta did not alter radiation sensitivity, it did increase resistance to cisplatin and induced resistance to hyperthermia at higher temperatures (45degreesC).

We investigated the ability of camptothecin to potentiate cell killing by low-dose-rate irradiation and whether this potentiation was associated with an increase in the level of residual DNA double-strand breaks (DSBs). Human melanoma (Sk-Mel-3) cells, grown to the confluent phase, were treated with low-dose-rate radiation (0.88 cGy/min) alone, camptothecin alone, or concurrent camptothecin and low-dose-rate radiation. Cell survival was determined using a clonogenic assay. The interactions between camptothecin and low-dose-rate radiation were analyzed further using isobolograms. DNA DSBs were determined using the neutral comet assay. We found that 10 and 25 muM camptothecin, but not 1 muM, camptothecin potentiated cell killing significantly relative to that seen with low-dose-rate radiation alone. Unexpectedly, the potentiation of the effects of low-dose-rate radiation by camptothecin was accompanied by large Increases in the alpha parameter of the linear-quadratic fit rather than in the beta parameter. This suggests a modification of intrinsic radiosensitivity rather than of repair of sublethal damage. From isobologram analysis, low-dose-rate radiation interacted either additively or supra-additively with 25 or 10 muM camptothecin. Conversely, the interaction of low-dose-rate radiation with 1 muM camptothecin was subadditive. Finally, there were strong correlations (correlation coefficients >0.9) between surviving fraction and either comet tail length or comet tail moment after concurrent treatment with 25 muM camptothecin and low-dose-rate radiation. This suggests that the level of residual DNA DSBs was a good indicator of cell killing after treatment with low-dose-rate radiation plus 25 muM camptothecin. (C) 2002 by Radiation Research Society.

To determine whether pulsed dose rate irradiation in combination with mild hyperthermia could radiosensitize cells in comparison to pulsed dose rate irradiation alone, human ovarian carcinoma (A2780s, cisplatin- and radiation-sensitive, and A2780cp, cisplatin- and radiation-resistant) and human fibroblast (AG1522) cell lines were used. Cells were irradiated in vitro using two fraction sizes, 0.53 Gy given every hour and 1.6 Gy given every 3h, with an overall average dose rate of 0.53 Gy/h. The data showed that 40 degrees C hyperthermia did not radiosensitize any of the cell lines for the 0.53 Gy every 1 h fractionation scheme. In addition, mild hyperthermia radiosensitized both carcinoma cell lines when using the 1.6 Gy fraction size for all doses tested in the A2780s and at higher doses in the A2780cp, but not the normal cell line. These results suggest a potential clinical advantage when using the 1.6 Gy fraction size with 40 degrees C mild hyperthermia, since hyperthermia radiosensitized the carcinoma cells but not the normal cells.