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Reirradiation (reRT) for locoregional recurrences poses unique challenges and risks; re-treatment using proton beam radiotherapy (PBT) could prove advantageous. Assessing clinical outcomes and toxicity profiles, this systematic review comprehensively evaluated available evidence regarding PBT reRT. Fourteen original investigations across central nervous system (CNS) (n=6), head/neck (H&N) (n=4), lung (n=2), and gastrointestinal (n=2) malignancies were analyzed. PBT for recurrent uveal melanoma achieved 5-year eye retention of 55%; for chordomas, reRT afforded a 2-year local control and overall survival (OS) of 85% and 80%, respectively. Multiple PBT reRT studies for adult gliomas illustrate no grade ≥3 toxicities. Two pediatric CNS tumor studies demonstrated the safety and efficacy of reRT, with one total grade 3 toxicity and achievement of longer-term OS. PBT for H&N malignancies shows appropriate local/locoregional control and favorable toxicity profiles versus historical photon-based methods, including low (9-10%) rates of feeding tube placement. PBT for recurrent lung cancer can achieve favorable survival with expected toxicities/complications of reRT, especially with concurrent chemotherapy and centrally located recurrences. Lastly, PBT reRT in gastrointestinal malignancies induced very few high-grade complications. Hence, based on the limited existing data, PBT is a notably safe reRT modality for effective salvage of recurrent disease. Institutional experiences must continue to be reported: dosimetric correlations, late toxicities, and advanced PBT techniques., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Purpose: We assessed clinical and radiologic outcomes in adults and children with low-grade glioma (LGG) of the brain treated with pencil-beam scanning (PBS) proton therapy (PT)., Materials and Methods: Between 1997 and 2014, 28 patients were treated with PBS PT, 20 (71%) of whom were younger than 18 years. Median age at start of PT was 12.3 years (range, 2.2-53.0 years). Nine patients (32%) underwent at least a subtotal resection; 12 (43%) underwent biopsy; and 7 (25%) were diagnosed radiographically. Twelve patients (43%) had grade II and 9 (32%) had grade I gliomas. Eleven patients (39%) received chemotherapy before PT. A median dose of 54 Gy (relative biologic effectiveness) was administered. Radiologic response to PT was determined using the Response Evaluation Criteria in Solid Tumors (RECIST). Eight domains of quality of life (QoL) for 16 pediatric patients were assessed prospectively by patients' parents using the pediatric QoL proxy questionnaire. Progression-free survival and overall survival (OS) were estimated by the Kaplan-Meier method. Median follow-up was 42.1 months for living patients., Results: Ten patients (36%) developed local, clinical failure. Three patients (11%) died, all of tumor progression. Radiographic tumor response by RECIST was evaluable in 11 patients: 9 (82%) with stable disease, 1 (9%) with partial response, and 1 (9%) with complete response to PT. Three-year OS and progression-free survival were 83.4% and 56.0%, respectively. No ≥ grade III acute toxicities were observed. Grade III, late radiation necrosis developed in 1 patient (4%). No appreciable change in pediatric QoL proxy scores in children was noted in any of the 8 domains at any time point., Conclusion: Treatment with PBS PT is effective for LGG, with minimal acute toxicity and, in children, no appreciable decline in QoL. More patients and longer follow-up are needed to determine the long-term efficacy and toxicity of PT for LGG., Competing Interests: Conflicts of Interest: The authors have no conflicts of interest to disclose., (© Copyright 2017 International Journal of Particle Therapy.)

Objectives: The aim of this study was to evaluate the long-term effectiveness and complications of radiotherapy (RT) in the treatment of patients with mucosal melanomas of the head and neck., Materials and Methods: The medical records of 21 patients treated with definitive or postoperative (RT) between 1974 and 2011 at the University of Florida Department of Radiation Oncology in Gainesville, FL, and the University of Florida Proton Therapy Institute in Jacksonville, FL, were retrospectively reviewed under an Institutional Review Board-approved protocol. Primary sites included nasal cavity, oropharynx, and paranasal sinuses. Sixteen patients (76%) received surgery and postoperative RT and 5 patients (24%) received RT alone. Seventeen patients received photon RT alone, whereas 4 patients received combined photon-based and proton-based RT. Median follow-up for all patients was 1.05 years (range, 0.36 to 12.97 y); median follow-up for survivors was 2.2 years (range 0.9 to 13.0 y)., Results: The 5-year outcomes were: local control, 79%; regional control, 85%; local-regional control, 65%; distant metastasis-free survival, 20%; cause-specific survival, 22%; and overall survival, 22%. Three patients (14%) experienced severe complications including bilateral blindness and skin necrosis., Conclusions: Definitive or postoperative RT for mucosal melanoma of the head and neck yields fairly good local-regional control of disease. The prognosis for patients treated with definitive RT is less promising than for those who receive surgery and postoperative RT.

Purpose: To evaluate the long-term effectiveness of radiotherapy (RT) in the treatment of sinonasal undifferentiated carcinoma (SNUC)., Materials and Methods: The medical records of 23 patients treated with definitive or postoperative RT between 1992 and 2010 at the University of Florida were retrospectively reviewed. Fifteen patients (65%) received primary surgery and postoperative RT. Radiation doses ranged from 59.0 to 74.8 Gy (median, 70.2 Gy). The median follow-up time for all patients was 3.0 years (range, 0.9-19.9), and for living patients was 7.7 years (range, 2.5-19.9)., Results: The actuarial 5-year survival outcomes were as follows: progression-free survival, 42%; cause-specific survival, 43%; and overall survival, 32%. Actuarial 5-year disease control rates were as follows: local control (infield or marginal), 74%; local-regional control (excluding leptomeningeal spread), 58%, regional control 78%, freedom from leptomeningeal recurrence, 72%, and distant metastasis-free survival, 73%. Five of the 8 (62.5%) patients treated with definitive RT died with disease, and 6 of the 15 patients (40%) treated with primary surgery and postoperative RT died with disease. Three patients (13%) experienced severe complications including unilateral eye removal, osteoradionecrosis of the maxilla requiring hyperbaric oxygen and surgery, and brain necrosis. One patient died due to an infected bone graft and brain abscess., Conclusions: A multimodal approach is best when treating SNUC patients. The prognosis for patients treated with definitive RT ± chemotherapy is less promising than for those who receive surgery and postoperative RT ± chemotherapy. Severe complications occur in about 17% of patients due to the high dose of RT alone or combined with surgery required for acceptable disease control., (© 2014.)

Objectives Skull base chordoma is a rare, locally aggressive tumor located adjacent to critical structures. Gross total resection is difficult to achieve, and proton therapy has the conformal advantage of delivering a high postoperative dose to the tumor bed. We present our experience using proton therapy to treat 33 patients with skull base chordomas. Design Retrospective outcomes study. Setting University of Florida Proton Therapy Institute; 2007 to 2011. Participants A total of 33 patients with skull base chordomas received postoperative three-dimensional conformal proton therapy. The patients were 79% male and 6% diabetic; 27% had received a gross total resection. Main Outcome Measures The gross tumor/tumor bed received a dose between 77.4 CGE and 79.4 CGE. Local control and overall survival were tracked, and radiation toxicity was assessed using a modified Radiation Therapy Oncology Group/European Organization for Research and Treatment of Cancer Late Radiation Morbidity Scoring Scheme. Results Median follow-up for all patients was 21 months. Local control and overall survival rates at 2 years were 86% and 92%, respectively. Grade 2 toxicity was observed in 18% of our cohort in the form of unilateral hearing loss partially corrected with a hearing aid. No grade 2 or higher optic or brainstem toxicities were observed. Conclusions Proton therapy is an effective treatment modality for skull base chordomas.

Objective: The objective of the study was to discuss the optimal management and treatment outcomes for patients with head and neck osteosarcomas., Study Design: Review article., Methods: Review of the pertinent literature., Results: Osteosarcomas account for approximately 1% or less of all head and neck cancers. The vast majority occur in the mandible and maxilla. The median age is in the fourth decade, with a wide range. They are more likely to recur locally after treatment and distant metastases are observed less often than with the more common osteosarcomas arising in the long bones. The optimal treatment is complete resection. The role of adjuvant chemotherapy is ill-defined. The vast majority of recurrences are observed within 5 years. The 5-year disease-specific and overall survival rates are approximately 60% to 70%., Conclusions: Osteosarcoma of the head and neck is a rare entity that occurs primarily in the mandible and maxilla. The optimal treatment is surgery. Adjuvant radiotherapy should be considered for those with close or positive margins. The role of adjuvant chemotherapy is ill-defined. The likelihood of cure is approximately 60% to 70%., (Copyright © 2011 Elsevier Inc. All rights reserved.)

There is a strong rationale for potential benefits from proton therapy (PT) for selected cancers of the head and neck because of the opportunity to improve the therapeutic ratio by improving radiation dose distributions and because of the significant differences in radiation dose distribution achievable with x-ray-based radiation therapy (RT) and PT. Comparisons of dose distributions between x-ray-based and PT plans in selected cases show specific benefits in dose distribution likely to translate into improved clinical outcomes. However, the use of PT in head and neck cancers requires special considerations in the simulation and treatment planning process, and currently available PT technology may not permit realization of the maximum potential benefits of PT. To date, few clinical data are available, but early clinical experiences in sinonasal tumors in particular suggest significant improvements in both disease control and radiation-related toxicity.

Purpose: Downstaging (DS) of rectal cancers is achieved in approximately 45% of patients with neoadjuvant fluorouracil (FU) -based chemoradiotherapy (CRT). Polymorphisms in the thymidylate synthase gene (TYMS) had previously defined two risk groups associated with disparate tumor DS rates (60% v 22%). We conducted a prospective single-institution phase II study using TYMS genotyping to direct neoadjuvant CRT for patients with rectal cancer., Patients and Methods: Patients with T3/T4, N0-2, M0-1 rectal adenocarcinoma were evaluated for germline TYMS genotyping. Patients with TYMS *2/*2, *2/*3, or *2/*4 (good risk) were treated with standard chemoradiotherapy using infusional FU at 225 mg/m²/d. Patients with TYMS *3/*3 or *3/*4 (poor risk) were treated with FU/RT plus weekly intravenous irinotecan at 50 mg/m². The primary end point was pathologic DS. Secondary end points included complete tumor response (ypT0), toxicity, recurrence rates, and overall survival., Results: Overall, 135 patients were enrolled, of whom 27.4% (37 of 135) were considered poor risk. The prespecified statistical goals were achieved, with DS and ypT0 rates reaching 64.4% and 20% for good-risk and 64.5% and 42% for poor-risk patients, respectively., Conclusion: To our knowledge, this is the first study to prospectively use TYMS genotyping to direct neoadjuvant CRT in patients with rectal cancer. High rates of DS and ypT0 were achieved among both risk groups when personalized treatment was based on TYMS genotype. These results are encouraging, and further evaluation of this genotype-based strategy using a randomized study design for locally advanced rectal cancer is warranted.

Objectives/hypothesis: To determine the outcomes after radiotherapy (RT) alone or combined with surgery at the University of Florida for patients with carcinomas of the nasal cavity and paranasal sinuses., Methods: Between November 1964 and June 2005, 109 patients were treated with curative intent. Patients with maxillary sinus carcinomas were excluded. Fifty-six patients were treated with definitive RT, and 53 patients received surgery and preoperative (eight patients) or postoperative (45 patients) RT. Median follow-up was 4.3 years (range, 0.2-35.9 years). Median follow-up on living patients was 9.4 years (range, 2.0-35.9 years)., Results: The 5-year local control rates were: T1-T3, 82%; T4, 50%; and overall, 63%. Local control at 5 years was 43% after definitive RT versus 84% after surgery and adjuvant RT (P < .0001). Multivariate analysis of local control revealed that both overall stage and treatment group (definitive RT versus surgery and adjuvant RT) significantly impacted this endpoint. Cause-specific survival rates were: stages I to III, 81%; stage IV, 54%; and overall, 62%. Multivariate analysis revealed that T-stage, N-stage, and treatment group significantly influenced this endpoint. Thirty-one (20%) of 109 patients sustained severe complications; 17 of 56 patients (16%) after definitive RT and 14 of 53 patients (25%) after surgery and adjuvant RT., Conclusions: The probability of local control and cause-specific survival is better after surgery and RT compared with definitive RT. There is a modest increase in the risk of complications after surgery and RT. Thus, the preferred treatment is surgery combined with preoperative or postoperative RT.

Objectives: To investigate the incidence of postradiation therapy (postRT) hypopituitarism (hp-pit) in pediatric patients treated for extracranial head and neck tumors., Methods: We retrospectively reviewed 30 pediatric patients treated with RT for extracranial head and neck tumors between 1970 and 2000. RT doses to the hypothalamic-pituitary axis were estimated by re-evaluating the treatment plans., Results: Clinical hp-pit was observed in 16 (53.3%) patients; the median detection latency was 3.3 years and the median dose to the pituitary was 40.7 Gy. Univariate analysis of clinical hp-pit revealed that the total dose to the pituitary was significant (P = 0.034) as was the patient's age at the time of RT (P = 0.018) showing higher susceptibility and more damage in younger patients. Though fractionation (P = 0.018) and adjuvant chemotherapy (P = 0.016) were significant in univariate analysis, the total dose received by patients in these 2 groups was higher than the median dose for all patients. A limited multivariate analysis with 3 variables indicated that the total dose to the pituitary was significant, but chemoradiation and fractionation were not. The 5- and 10-year rates of freedom from clinical hp-pit were 42% and 37%, respectively., Conclusions: Clinical manifestations of late radiation toxicity to the hypothalamic-pituitary axis were observed among the pediatric patients with hormone deficiencies. The development of hp-pit depends on the total dose received by the hypothalamic-pituitary axis and the age of the patient. Dependence on chemoradiation and fractionation were inconclusive. Relatively long latency before clinical manifestations indicates that periodic testing after RT in pediatric patients is necessary for early detection and management of hormonal deficiencies.

Purpose: The response of rectal cancers to neoadjuvant chemoradiotherapy is variable. Tumor hypoxia reduces the effectiveness of both radiation therapy and chemotherapy and is a well-known risk factor for tumor radioresistence. We hypothesized that imaging with the novel hypoxia-detecting agent, (60)Cu-diacetyl-bis (N(4)-methylthiosemicarbazone) ((60)Cu-ATSM), previously validated in cervical and lung cancers, would predict the response of rectal cancers to neoadjuvant chemoradiotherapy and prognosis., Methods: Patients with locally invasive (T2-4) primary or node-positive rectal cancer located <12 cm from the anal verge were recruited for this pilot study. Pretreatment tumor size and stage were determined by endorectal ultrasonography, CT, and magnetic resonance imaging. Eleven patients also underwent clinical positron emission tomography with (18)F-fluorodeoxyglucose at the discretion of the treating clinician. The primary tumor was imaged by positron emission tomography with (60)Cu-ATSM, and accumulation of the tracer was measured semiquantitatively by determining the tumor-to-muscle activity ratio. Neoadjuvant chemoradiotherapy was then administered (within 2 weeks of (60)Cu-ATSM-positron emission tomography) and consisted of 45 Gy given in 25 fractions to the pelvis with continuous intravenous infusion of 5-fluorouracil (225 mg/m(2)/day). Proctectomy was performed six to eight weeks after neoadjuvant chemoradiotherapy and the tumor submitted to pathology for size measurement and staging. Tumor-to-muscle activity ratios were compared with tumor (18)F-fluorodeoxyglucose uptake, tumor response to neoadjuvant chemoradiotherapy, and with patient survival., Results: Nineteen patients were enrolled in the study, two of whom were excluded from final analysis (1 death during neoadjuvant chemoradiotherapy and 1 tumor perforation during neoadjuvant chemoradiotherapy requiring emergent surgery). Of the 17 remaining patients, 14 had a reduction in tumor size and 13 were downstaged. The median tumor-to-muscle activity ratio of 2.6 discriminated those with worse prognosis from those with better prognosis. Both overall and progression-free survivals were worse with hypoxic tumors (tumor-to-muscle activity ratio >2.6) than with nonhypoxic tumors (tumor-to-muscle activity ratio 2.6 (positive predictive value 66 percent), whereas 6 of 14 with decreased size had tumor-to-muscle activity ratios >2.6 (negative predictive value 57 percent). Three of the 4 tumors not downstaged had tumor-to-muscle activity ratios >2.6 (positive predictive value 75 percent), whereas 5 of 13 downstaged tumors had tumor-to-muscle activity ratios >2.6 (negative predictive value 62 percent). The mean tumor-to-muscle activity ratio for downstaged tumors (2.2) was significantly lower than that of nondownstaged tumors (3.3) (P = 0.03). The difference in mean tumor-to-muscle activity ratio between downsized (2.3) and nondownsized (2.9) tumors did not reach statistical significance (P = 0.36). Tumor (18)F-fluorodeoxyglucose uptake (n = 11) did not correlate with (60)Cu-ATSM uptake (r = 0.4; P = 0.9) and there was no significant difference in mean tumor (18)F-fluorodeoxyglucose uptake between patients with hypoxic tumors and those with normoxic tumors (P = 0.3)., Conclusions: The results of this small pilot study suggest that (60)Cu-ATSM-PET may be predictive of survival and, possibly, tumor response to neoadjuvant chemoradiotherapy in patients with rectal cancer. A larger Phase II study is warranted to validate these results.

Background: The purpose of this study was to investigate the incidence of radiotherapy-induced primary and secondary hypopituitarism., Methods: Three hundred twelve patients treated with radiotherapy for extracranial head and neck tumors between 1964 and 2000 were evaluated. Radiotherapy doses to the hypothalamus and pituitary were estimated by reconstructing treatment plans., Results: Clinical hypopituitarism was observed in 44 (14.1%) patients after a median interval of 5.6 years. Hypothalamic dysfunction was reported in 14 patients after a median interval of 4.4 years. Neither fractionation nor adjuvant chemotherapy significantly impacted clinical hypopituitarism on multivariate analysis, but total dose to the pituitary was significant (p = .0228). Twenty-three of 68 (33.8%) patients tested for hypopituitarism demonstrated subclinical hypopituitarism. The 5- and 10-year rates of freedom from clinical hypopituitarism were 93% and 72%. The 5- and 10-year rates of freedom from subclinical hypopituitarism were 85% and 65%., Conclusion: Clinical and subclinical manifestations of late radiation toxicity were observed in the hypothalamic-pituitary axis.

To determine the efficacy of re-irradiation of patients with head and neck carcinoma we reviewed the pertinent literature. Depending on the location and extent of the tumor, re-irradiation may be accomplished with external beam radiotherapy (RT), brachytherapy, intraoperative RT, and/or radiosurgery. The likelihood of cure is impacted by the interval between the initial course of RT and re-irradiation, whether the carcinoma is a recurrence or a second primary tumor, initial T-stage, and recurrent T-stage (rT-stage), whether the gross disease is isolated or local-regional, and histology. Patients with T1-T2 N0 second primary tumors have a relatively high likelihood of cure, whereas those with recurrent T3-T4 cancers, a short disease-free interval, and/or associated with a regional recurrence have a very low probability of cure. The likelihood of severe late complications is relatively high and is related to prior RT dose, primary site, retreatment RT dose, treatment volume, and technique. Previously irradiated patients with T1-T2 N0 carcinomas who are not surgical candidates may benefit from re-irradiation, albeit with an elevated risk of late complications. Those with advanced recurrences are unlikely to benefit from re-irradiation and should be considered for either a prospective clinical trial or for palliative chemotherapy or supportive care.

Purpose: This is a phase II, single-center, single-arm study of patients with resectable adenocarcinoma of the pancreas who were treated with adjuvant interferon-based chemoradiation followed by gemcitabine. The primary end point was 2-year overall survival, with secondary endpoints being 2-year disease-free survival, and the frequency of grade 3 or 4 toxicity., Patients and Methods: From April 2002 to September 2005, 53 patients with adenocarcinoma of the pancreas underwent curative resection at a single institution, and subsequently received interferon- and gemcitabine-based adjuvant therapy consisting of external-beam irradiation at a dose of 5040 cGy (25 fractions per 5 weeks) and simultaneous 3-drug chemotherapy consisting of (1) continuous infusion 5-fluorouracil (175 mg/m2); (2) weekly intravenous bolus cisplatin (25 mg/m2); and (3) interferon-alpha (3 million units subcutaneously 3 times per week) during the 6 weeks of radiation. This was followed by two 4-week courses of weekly intravenous infusion of gemcitabine (1000 mg/m2, 3 of 4 weeks)., Results: Median follow-up is 38 months. Seventy-seven percent of patients had node-positive disease. Sixteen patients (30%) failed to complete adjuvant therapy, due to disease progression (7 patients), toxicity (7 patients), and consent withdrawal (2 patients). No patients completed planned therapy without dose modification. Median overall survival was 25 months (confidence interval [CI] = 21.5-48.5 months). Actuarial overall survival for the 1-, 2- and 3-year periods were 75% (CI = 61-85%), 56% (CI = 41-69%), and 41% (26-55%), respectively., Conclusions: This phase II trial demonstrated increased patient survival compared with historical controls, and equivalent survival compared with the regimen combining interferon-alpha with 5-fluorouracil-based chemoradiation. Despite these encouraging results, significant concerns regarding dose- and treatment-limiting toxicities remain.

We discuss the optimal treatment and outcomes for pleomorphic adenoma of the salivary glands by reviewing the pertinent literature. Pleomorphic adenoma is the most common benign salivary gland neoplasm. It is found mostly in the parotid gland in middle-aged women. It progresses slowly and, left untreated, can produce significant morbidity and, rarely, death. The optimal treatment is superficial or total parotidectomy with facial nerve preservation, which results in local control rates of 95% or higher. Radiotherapy (RT) is useful to obtain local control in patients with positive margins, unresectable tumors, and multifocal recurrences after prior resection. Local control rates after RT for microscopic and gross residual tumor are approximately 80% to 85% and 40% to 60%, respectively. The main complication is surgically induced 7th nerve injury. Surgery is the mainstay of treatment and results in a very high cure rate. RT increases the likelihood of local control in the small subset of patients with incompletely resectable tumors and/or multifocal recurrences.

The purpose of this article is to review the pertinent literature and discuss the optimal treatment and outcomes for patients with ameloblastoma. Ameloblastoma is an uncommon benign, locally aggressive odontogenic neoplasm that usually occurs in the vicinity of the mandibular molars or ramus. Uncontrolled, ameloblastoma may cause significant morbidity and occasionally death. The median age is approximately 35 years and males and females are equally affected. The majority of ameloblastomas are multicystic, which are more difficult to eradicate than the unicystic and peripheral varieties. Although surgery is the mainstay of treatment, the extent of resection is controversial. Radical resections, including marginal and segmental mandibulectomy, result in local control rates exceeding 90%. In contrast, conservative procedures such as enucleation and/or curettage result in local control rates of approximately 80% and 50% for unicystic and multicystic ameloblastomas, respectively. Limited experience with radiotherapy indicates that it may reduce the risk of progression and result in long-term local control in the occasional patient with incompletely resectable disease. The optimal treatment for ameloblastoma is wide en bloc resection. Radiotherapy may improve the likelihood of local control in the occasional patient with incompletely resectable tumor.

Purpose: To discuss the treatment and outcomes of inverted papilloma of the nasal cavity and paranasal sinuses., Methods: Review of the pertinent literature., Results: Inverted papilloma is a benign, locally aggressive neoplasm that arises in the nasal cavity and is associated with squamous cell carcinoma in approximately 5% of patients. Squamous cell carcinoma may be present with inverted papilloma at the initial diagnosis or it may occur metachronously after prior treatment. Surgery is the primary treatment of inverted papilloma. The likelihood of local recurrence varies from less than 5% to over 50%, depending on the extent of resection. There is likely no significant difference in the risk of local recurrence after open compared with endoscopic resection. The probability of local recurrence and/or death from tumor is increased if inverted papilloma is associated with squamous cell carcinoma. The likelihood of cure is approximately 50% when malignancy is present and postoperative radiotherapy should be considered for the majority of patients. A small subset of patients with inverted papilloma present with incompletely resectable disease. Definitive radiotherapy using doses between 65 and 70 Gy will locally control gross disease in the majority of patients., Conclusion: The preferred treatment of inverted papilloma is surgery; postoperative radiotherapy is added if it is associated with squamous cell carcinoma. The likelihood of local recurrence after surgery for inverted papilloma may be substantial and varies with the extent of resection. Definitive radiotherapy may be used to successfully treat patients with incompletely resectable inverted papilloma.

Purpose: To determine if hypoxia-related molecular markers are associated with (60)Cu labeled diacetyl-bis (N4 -methylthiosemicarbazone); ((60)Cu-ATSM) imaging of tumor hypoxia in cervical cancer., Procedures: Fifteen patients were enrolled in a prospective study and underwent evaluation of tumor hypoxia with positron emission tomography (PET) using (60)Cu-ATSM. (60)Cu-ATSM-PET imaging was compared with the expression of tissue molecular markers, which included vascular endothelial growth factor (VEGF), cyclo-oxygenase-2 (COX-2), epidermal growth factor receptor (EGFR), carbonic anyhdrase IX (CA-9), and apoptotic index., Results: Six patients had hypoxic tumors determined by (60)Cu-ATSM, and nine had non-hypoxic tumors. The 4-year overall survival estimates were 75% for patients with non-hypoxic tumors and 33% for those with hypoxic tumors (p = 0.04). Overexpression of VEGF (p = 0.13), EGFR (p = 0.05), CA-9 (p = 0.02), COX-2 (p = 0.08), and the presence of apoptosis (p = 0.005) occurred in patients with hypoxic tumors. Cox proportional hazards modeling demonstrated hypoxia as determined by (60)Cu-ATSM to be a significant independent predictor of tumor recurrence (p = 0.0287)., Conclusions: (60)Cu-ATSM hypoxia was correlated with overexpression of VEGF, EGFR, COX-2, CA-9, an increase in apoptosis, and a poor outcome.

Purpose: To investigate the incidence of radiotherapy (RT)-induced central and primary hypothyroidism regarding total dose, fractionation, and adjuvant chemotherapy., Methods and Materials: We retrospectively reviewed the data from 312 patients treated with RT for extracranial head-and-neck tumors between 1964 and 2000. The cervical lymph nodes were irradiated in 197 patients. The radiation doses to the thyroid gland and hypothalamic-pituitary axis were estimated by reconstructing the treatment plans., Results: Clinical central hypothyroidism (CH) was observed in 17 patients (5.4%); the median clinical latency was 4.8 years. Clinical primary hypothyroidism (PH) was observed in 40 patients (20.3%); the median clinical latency was 3.1 years. Multivariate analysis of clinical CH revealed that fractionation, adjuvant chemotherapy, and total dose to the pituitary were not significant. Multivariate analysis of clinical PH revealed that the total dose to the thyroid (p = 0.043) was significant, but adjuvant chemotherapy, age, and gender were not. Of the patients tested for hypopituitarism, 14 (20.3%) of 69 demonstrated subclinical CH and 17 (27.4%) of 62 demonstrated subclinical PH. The 5-year and 10-year rates of freedom from clinical CH and PH were 97% and 87% and 68% and 67%, respectively. Of the patients tested, the 5-year and 10-year rates of freedom from subclinical CH and PH were 91% and 78% and 71% and 71%, respectively., Conclusion: Clinical and subclinical manifestations of late radiation toxicity were observed in the thyroid and hypothalamic-pituitary axis. Although CH did not indicate a dependence on fractionation, adjuvant chemotherapy, or total dose to the pituitary, PH showed a dependence on the total dose to the thyroid gland.

Perineural invasion occurs in 2% to 6% of cutaneous basal and squamous cell carcinomas of the head and neck and is associated with midface location, recurrent tumors, high histologic grade, and increasing tumor size. Patients may be asymptomatic with perineural invasion appreciated on pathologic examination of the surgical specimen (incidental) or may present with cranial nerve deficits (clinical). The cranial nerves most commonly involved are the 5th and 7th nerves. Magnetic resonance imaging is obtained to detect and define the extent of perineural invasion; computed tomography is used to detect regional lymph node metastases. Patients with apparently resectable cancers undergo surgery usually followed by postoperative radiotherapy. Patients with incompletely resectable cancers are treated with definitive radiotherapy. The 5-year local control, cause-specific survival, and overall survival rates are approximately 87%, 65%, and 50%, respectively, for patients with incidental perineural invasion compared with 55%, 59%, and 55%, respectively, for those with clinical perineural invasion.

The management of head and neck cancer has evolved into a multidisciplinary approach in which patients are evaluated before treatment and decisions depend on prospective multi-institutional trials, as well as retrospective outcome studies. The choice of one or more modalities to use in a given case varies with the tumor site and extent, as exemplified in the treatment of laryngeal squamous cell carcinomas. The goals of treatment include cure, laryngeal voice preservation, voice quality, optimal swallowing, and minimal xerostomia. Treatment options include transoral laser excision, radiotherapy (both definitive and postoperative), open partial laryngectomy, total laryngectomy, and neck dissection. The likelihood of local control and preservation of laryngeal function is related to tumor volume. Patients who have a relatively high risk of local recurrence undergo follow-up computed tomography scans every 3-4 months for the first 2 years after radiotherapy. Patients with suspicious findings on computed tomography might benefit from fluorodeoxyglucose positron emission tomography to differentiate post-radiotherapy changes from tumor.

Purpose: We sought to report the outcomes after definitive radiotherapy (RT) for nasopharyngeal carcinoma., Methods: Between March 1983 and November 2003, 82 patients were treated with curative intent and followed from 0.2 to 22 years (median, 5 years). Follow-up on living patients ranged from 2.8 to 22 years (median, 10.8 years). Thirty-two patients (39%) were treated once daily, 45 patients (55%) were treated with hyperfractionation, and 5 patients (6%) were treated with the concomitant boost technique. Eight patients (10%) received intensity-modulated radiotherapy. Thirty-one patients (38%) received induction (17 patients) or concomitant (14 patients) chemotherapy. Fourteen patients (17%) underwent a planned neck dissection., Results: The 5-year outcomes after treatment were: local control, 78%; regional control, 90%; local-regional control, 76%; distant metastasis-free survival, 80%; cause-specific survival, 66%; and survival, 57%. The impact of T-stage, N-stage, overall stage, World Health Organization class, fractionation schedule, and adjuvant chemotherapy were evaluated in a multivariate analysis of various outcomes. Only N-stage and World Health Organization class impacted regional control., Conclusions: The likelihood of cure after RT is relatively high. Although the likelihood of cure diminished with increasing stage, none of the parameters evaluated significantly influenced outcomes aside from N-stage and WHO class.

This review discusses the role of postoperative radiotherapy (RT) for patients with squamous cell carcinoma of the head and neck. Patients with unfavorable pathologic features have a high-risk of local-regional recurrence and a decreased likelihood of survival after surgery alone. Postoperative RT reduces the risk of local-regional failure and probably improves survival. Patients who are at high risk for recurrence may benefit from more aggressive altered fractionation schedules to decrease the overall time from surgery to the completion of RT. Adjuvant cisplatin-based chemotherapy also appears to improve the probability of cure in high-risk patients.

Purpose: Surgical staging and treatment of anal carcinoma has been replaced by noninvasive staging studies and combined modality therapy. In this study, we compare computed tomography (CT) and physical examination to [(18)F]-fluoro-2-deoxy-D-glucose-positron emission tomography/computed tomography (FDG-PET/CT) in the staging of carcinoma of the anal canal, with special emphasis on determination of spread to inguinal lymph nodes., Methods and Materials: Between July 2003 and July 2005, 41 consecutive patients with biopsy-proved anal carcinoma underwent a complete staging evaluation including physical examination, CT, and 2-FDG-PET/CT. Patients ranged in age from 30 to 89 years. Nine men were HIV-positive. Treatment was with standard Nigro regimen., Results: [(18)F]-fluoro-2-deoxy-D-glucose-positron emission tomography/computed tomography (FDG-PET/CT) detected 91% of nonexcised primary tumors, whereas CT visualized 59%. FDG-PET/CT detected abnormal uptake in pelvic nodes of 5 patients with normal pelvic CT scans. FDG-PET/CT detected abnormal nodes in 20% of groins that were normal by CT, and in 23% without abnormality on physical examination. Furthermore, 17% of groins negative by both CT and physical examination showed abnormal uptake on FDG-PET/CT. HIV-positive patients had an increased frequency of PET-positive lymph nodes., Conclusion: [(18)F]-fluoro-2-deoxy-D-glucose-positron emission tomography/computed tomography detects the primary tumor more often than CT. FDG-PET/CT detects substantially more abnormal inguinal lymph nodes than are identified by standard clinical staging with CT and physical examination.

Purpose: The purpose of this study is to update our experience with definitive radiotherapy (RT) for carcinoma of the tonsillar area., Patients and Methods: There were 503 patients treated between October 1964 and May 2003 (potential follow-up for at least 2 years). Of these, 198 patients underwent a planned neck dissection and 57 patients received induction (18 patients) or concomitant (39 patients) chemotherapy., Results: The 5-year local control rates were as follows: T1, 88%; T2, 84%; T3, 78%; and T4, 61%. Multivariate analysis revealed that local control was significantly influenced by T stage, primary site, and fractionation. Local control after RT for early stage cancers was higher for tonsillar fossa/posterior pillar tumors than for those arising from the anterior tonsillar pillar. The 5-year cause-specific survival rates were as follows: I, 100%; II, 86%; III, 84%; IVA, 73%; and IVB, 46%. Multivariate analysis revealed that cause-specific survival was significantly influenced by T stage, overall stage, neck dissection, race, and gender. The incidence of severe late complications was 9%., Conclusion: Based on our data and a review of the literature, definitive RT provides cure rates that are as good as those after surgery, and is associated with a lower rate of severe complications. Patients with lateralized tumors may be safely treated with ipsilateral field arrangements. Our limited experience with intensity modulated radiotherapy suggests that it is as efficacious as conventional RT.

This paper examined heat-induced radiosensitization in two Chinese hamster heat-resistant cell lines, HR-1 and OC-14, that were isolated from the same wild-type HA-1 cell line. It found a reduction of the magnitude of heat-induced radiosensitization after exposure to 43 degrees C in both HR-1 and OC-14 cells and a similar reduction after exposure to 45 degrees C in HR-1 cells, but not in OC-14 cells. The effect of heat exposure on a class of ionizing radiation-induced DNA damage that inhibits the ability of nuclear DNA to undergo super-coiling changes was also studied using the fluorescent halo assay in these three cell lines. Wild type cells exposed to either 43 or 45 degrees C before irradiation had a DNA rewinding ability that was intermediate between control and unheated cells, a phenomenon previously described as a masking effect. This masking effect was significantly reduced in HR-1 cells exposed to either 43 or 45 degrees C or in OC-14 cells exposed to 43 degrees C under conditions that heat-induced radiosensitization was reduced. In contrast, the masking effect was not altered in OC-14 cells exposed to 45 degrees C, conditions under which heat-induced radiosensitization was similar to that observed in wild-type HA-1 cells. These results suggest that a reduction in the masking effect is associated with a reduction of the magnitude of heat-induced radiosensitization in the HR-1 and OC-14 heat-resistant cell lines. The reduction of the masking effect in the cell lines resistant to heat-induced radiosensitization was associated with neither a reduction in the magnitude of the heat-induced increase in total nuclear protein content nor major differences in the protein profiles of the nucleoids isolated from heated cells.

Purpose: To evaluate the utility of sequential (18)F-fluorodeoxyglucose positron emission tomography (FDG-PET) imaging for brachytherapy treatment planning in patients with carcinoma of the cervix., Methods and Materials: Twenty-four patients with carcinoma of the cervix were included in this prospective study. The clinical stage of their disease was Ib (7), IIa (1), IIb (7), and IIIb (9). Patients were treated with irradiation and brachytherapy, with the majority receiving concurrent weekly cisplatin chemotherapy. Patients underwent diagnostic FDG-PET imaging before treatment, sequential FDG-PET brachytherapy imaging during treatment, and diagnostic FDG-PET 3 months after treatment completion. Delineation of the gross tumor volume, bladder, and rectum was performed for all scans using a commercially available treatment-planning system. Actual treatment delivery was based on two-dimensional orthogonal planning., Results: The mean gross tumor volume and percent coverage by the target isodose surface for the initial, mid, and last implant were 37 cm3, 17 cm3, and 10 cm3 and 68%, 76%, and 79%, respectively. Nine of 11 patients were found to have continued decrease in tumor volume as measured by FDG-PET, with 3 patients having complete regression of their tumor before treatment was completed. The maximal bladder and rectal doses obtained from three-dimensional dose-volume histograms were significantly higher than the International Commission on Radiation Units and Measurements Report 38 bladder and rectal points obtained by two-dimensional treatment-planning., Conclusions: Sequential FDG-PET brachytherapy imaging identifies the tumor response in individual patients, potentially making patient-specific brachytherapy treatment planning possible.

Seventy-three patients were evaluated for the changes in pain relief, numbness and paresthesias after Gamma Knife radiosurgery to a maximum dose of 76-87 Gy for trigeminal neuralgia. Patients experienced pain relief as follows: 59% attained complete pain relief without prior surgery (33% with prior surgery); 25% achieved > or = 50% pain reduction (28% with prior surgery); 11% of surgery patients obtained minor pain relief, and 16% of patients without surgery had no relief (28% with prior surgery). Level of pain decreased rapidly within 6 weeks after radiosurgery. Numbness/paresthesias developed slowly over the first 12-15 months. Bothersome levels were experienced by 15% of the patients without prior surgery (22% with prior surgery). Comparison of the occurrence of numbness/paresthesias, with respect to prior surgery, was not statistically significant. Only 2% of all patients had persistently bothersome side effects. In conclusion, radiosurgery is an effective treatment of trigeminal neuralgia, especially for those patients not having prior surgery., (Copyright 2005 S. Karger AG, Basel.)

The customization of brachytherapy dose distributions for gynecologic malignancies is limited by the spatial positioning of the applicators. We tested the hypothesis that applicator-guided intensity modulated radiation therapy (AGIMRT) has the potential to deliver highly conformal dose distributions to cervical tumors, representing improvement over distributions obtained with intracavitary brachytherapy. A commercial three-dimensional (3-D) treatment planning system was used to create plans for ten cervical cancer patients treated at our institution. Dose distributions of conventionally designed high dose rate (HDR) plans were compared against those of AGIMRT. Tumor delineation was based on a previously published binary threshold technique, using image intensity on positron emission tomography (PET) scans. AGIMRT treatment schedules were designed using two fraction sizes: 6.5 Gy, to directly reproduce the HDR fractionation, and 1.8 Gy, to simulate traditional external beam fractionation. The average minimum tumor dose was significantly greater for the AGIMRT dose distributions than for the HDR distributions (64.2 Gy vs 33.6 Gy; p = 0.005). The mean percent tumor volume at the prescription dose was higher for the AGIMRT plans (90.0% vs 58.2%; p = 0.005). Using AGIMRT, the mean percent volume at the tolerance limit was decreased for the bladder (6.1% vs 16.6%; p = 0.047) but increased for the rectum (4.1% vs 2.2%; p = 0.646). Our study suggests that there may be conceptual and dosimetric advantages to replacing HDR with AGIMRT for patients with large-volume cervical tumors. This investigation is being expanded using sequential PET images to model tumor regression and compare brachytherapy and AGIMRT throughout the course of therapy.

Purpose: Computed tomography (CT)/positron emission tomography (PET)-guided intensity-modulated radiotherapy of the paraaortic lymph nodes (PALNs) has been proposed for patients with cervical carcinoma and paraaortic metastasis. This investigation attempted to determine the guidelines regarding the selection of appropriate treatment parameters (e.g., number of beams, beam geometry) and organ-specific parameters (e.g., importance weighting and tolerance dose) for intensity-modulated radiotherapy planning for the PALNs., Methods and Materials: Patients underwent imaging using CT and PET. The images were registered, and the structures were contoured. A goal dose of 50.4 Gy and 59.4 Gy was assigned to the clinical target volume (lymph node bed) and gross tumor volume (PET-delineated PALNs), respectively. For each patient, multiple treatment plans using various beam geometries and planning parameters were executed and evaluated in terms of the dose-volume histograms of the target and critical structures., Results: Acceptable sparing of the stomach, liver, and colon was achieved, regardless of the number of beams used. Sparing of the spinal cord was strongly dependent on the number and arrangement of the beams. Varying the number and arrangement of the beams affected small intestine sparing, but the amount of sparing was limited because the small intestine overlapped the target volumes, and, therefore, received the prescription dose. Adjusting the number of beams, beam angles, and prescription parameters provided minimal improvement in kidney sparing., Conclusion: We successfully developed treatment plans that deliver 59.4 Gy to the positive PALNs and 50.4 Gy to the paraaortic region using CT/PET-guided intensity-modulated radiotherapy.

Purpose: To evaluate a treatment planning method for dose escalation to the para-aortic lymph nodes (PALNs) based on positron emission tomography (PET) with intensity-modulated radiotherapy (IMRT) for cervical cancer patients with PALN involvement. One goal of this process was not to modify the traditional treatment of the pelvic region., Methods and Materials: PET images for 4 cervical cancer patents with PALN involvement were registered with their corresponding CT scans. Positive PALNs were identified on PET images, and the surrounding critical structures were delineated on CT images. The treatment machine central axis (CAX) was placed at the level of the L4-L5 vertebral body interspace. There were two distinct treatment regions: the para-aortic bed superior to the CAX and the whole pelvis region inferior to the CAX. IMRT was used for treatment planning of PALN bed irradiation. The positive PALNs identified on PET images were defined as the gross target volume, and the para-aortic bed was defined as the clinical target volume. The radiation doses were escalated from the conventional 45 Gy to 59.4 Gy for the gross target volume and 50.4 Gy for the clinical target volume in 33 fractions. The pelvis area was treated with conventional treatment methods, AP-PA beams to 50.4 Gy in 28 fractions with a brachytherapy implant boost. The placement of the CAX allowed the two treatment regions to be abutted using the treatment machine's independent jaws., Results: Dose escalation to positive PALNs, as identified on PET images, and the PALN bed is feasible with IMRT. Treatment plans for 4 patients revealed that escalated prescription doses could be delivered to target volumes while maintaining acceptable doses to the surrounding critical structures. Strategic placement of the treatment isocenter allows the IMRT region (PALN bed) and whole pelvis fields to be treated with a relatively uniform dose distribution in the abutment region., Conclusion: This study indicates that PET-guided IMRT could be used in a clinical trial in an attempt to escalate doses delivered to patients with cervical cancer who have positive PALNs.

Purpose: To compare conventional two-dimensional (2D) orthogonal radiography-based brachytherapy treatment planning for cervical cancer with a three-dimensional (3D) treatment planning technique based on 18F-fluoro-deoxyglucose-positron emission tomography (FDG-PET)., Methods and Materials: Eleven cervical cancer patients were included in this prospective study that evaluated one tandem and ovoid brachytherapy procedure for each patient. The patient underwent FDG-PET of the pelvis to visualize the tumor followed by a second FDG-PET scan with the FDG isotope placed inside the tandem and ovoid applicators to visualize the treatment source positions for 3D treatment planning. The tumor volumes were delineated using a binary threshold technique in which the threshold FDG-PET image intensity was 40% of the peak tumor intensity., Results: FDG-PET provides a reliable estimate of the cervical cancer volume and 3D spatial relationship of the tumor to the tandem and ovoid applicators. The maximal bladder and rectal doses determined from the 3D FDG-PET dose-volume histograms were found to be higher than those obtained using 2D treatment planning. The minimal dose to the tumor volume defined by FDG-PET ranged from 50 to 475 cGy for treatment plans designed to deliver 650 cGy to Point A and exhibited an inverse correlation with tumor volume., Conclusion: Physiologic FDG-PET brachytherapy treatment planning is feasible and accurate relative to conventional 2D treatment planning. The use of FDG-PET offers a unique method for tumor visualization and identifies the limitations of conventional brachytherapy treatment planning for coverage of large tumors and estimation of the dose to normal structures. This technique has the potential for improving isodose tumor coverage for patients with cervical cancer while sparing critical structures.

Purpose: Endocavitary radiation (RT) provides a conservative alternative to proctectomy. Although most suitable for small, mobile lesions, patients with less favorable tumors are often referred if they are poor surgical candidates. Knowing the extent to which radiation can control such tumors can be an important factor in making clinical decisions., Methods and Materials: One hundred ninety-nine patients, who received endocavitary RT with or without external beam RT (EBRT) during 1981 through 1995, were followed for disease status for a median of 70 months, including deaths from intercurrent causes. In the early years of the study, 21 patients were treated with endocavitary RT alone, the remainder of the patients received pelvic EBRT (usually 45 Gy in 25 fractions) 5-7 weeks before endocavitary RT., Results: Overall, 141 patients (71%) had local control with RT alone. Salvage surgery rendered an additional 20 patients disease free, for an ultimate local control rate of 81%. On multivariate analysis for local control (excluding surgical salvage), the most significant factors were mobility to palpation, use of EBRT, and whether pretreatment debulking of all macroscopic disease had been done (generally a piecemeal, nontransmural procedure). Of 77 cases staged by transrectal ultrasonography, the local control rate with RT alone was 100% for uT1 lesions, 85% (90% with no evidence of disease after salvage) for freely mobile uT2 lesions, and 56% (67% with no evidence of disease after salvage) for uT3 lesions and uT2 lesions that were not freely mobile., Conclusions: Patients with small mobile tumors that are either uT1 or have only a scar after debulking achieve excellent local control with endocavitary RT. About 15% of mobile uT2 tumors fail RT; therefore, careful follow-up is critical. Small uT3 tumors are appropriate for this treatment only if substantial contraindications to proctectomy are present.

Four cytometric assays for the assessment of radiation-induced DNA damage in individual cells are presented. Two of these, the alkaline and neutral comet assays, are useful for the detection of DNA damage due to very low radiation doses and promise to be useful for the quantitation of genomic damage after clinically or environmentally relevant exposures. The other two, the halo and halo-comet assays, reveal aspects of chromatin structure in the presence of DNA damage that reflect differences in intrinsic cellular radiosensitivity. Further development of these assays used alone, or in combination, should eventually lead to the definition of readily measurable cytometric parameters that will be useful as predictive markers for cellular responses to DNA damaging agents.

The effect of radiofrequency (RF) radiation in the cellular phone communication range (835.62 MHz frequency division multiple access, FDMA; 847.74 MHz code division multiple access, CDMA) on neoplastic transformation frequency was measured using the in vitro C3H 10T(1/2) cell transformation assay system. To determine if 835.62 MHz FDMA or 847.74 MHz CDMA radiations have any genotoxic effects that induce neoplastic transformation, C3H 10T(1/2) cells were exposed at 37 degrees C to either of the above radiations [each at a specific absorption rate (SAR) of 0.6 W/kg] or sham-exposed at the same time for 7 days. After the culture medium was changed, the cultures were transferred to incubators and refed with fresh growth medium every 7 days. After 42 days, the cells were fixed and stained with Giemsa, and transformed foci were scored. To determine if exposure to 835.62 MHz FDMA or 847.74 MHz CDMA radiation has any epigenetic effects that can promote neoplastic transformation, cells were first exposed to 4.5 Gy of X rays to induce the transformation process and then exposed to the above radiations (SAR = 0.6 W/kg) in temperature-controlled irradiators with weekly refeeding for 42 days. After both the 7-day RF exposure and the 42-day RF exposure after X irradiation, no statistically significant differences in the transformation frequencies were observed between incubator controls, the sham-exposed (maintained in irradiators without power to the antenna), and the 835.62 MHz FDMA or 847.74 MHz CDMA-exposed groups.

There have been allegations in the media and in the courts that cell phones and other types of hand-held transceivers are a cause of cancer. There have also been numerous public objections to the siting of TV, radio and cell phone transmission facilities because of a fear of cancer induction. A recent publication in Radiation Research by Repacholi et al. (147, 631-640, 1997) which suggests that exposure to radiofrequency (RF) radiation may increase lymphoma incidence in mice has contributed to this controversy. The goal of this review is to provide biomedical researchers a brief overview of the existing RF radiation-cancer studies. This article begins with a brief review of the physics and technology of cell phones. It then reviews the existing epidemiological studies of RF radiation, identifying gaps in our knowledge. Finally, the review discusses the cytogenetics literature on RF radiation and the whole-animal RF-radiation carcinogenesis studies. The epidemiological evidence for an association between RF radiation and cancer is found to be weak and inconsistent, the laboratory studies generally do not suggest that cell phone RF radiation has genotoxic or epigenetic activity, and a cell phone RF radiation-cancer connection is found to be physically implausible. Overall, the existing evidence for a causal relationship between RF radiation from cell phones and cancer is found to be weak to nonexistent.

The nuclear matrix organizes nuclear DNA into operational domains in which DNA is undergoing replication, transcription or is inactive. The proteins of the nuclear matrix are among the most thermal labile proteins in the cell, undergoing denaturation at temperatures as low as 43-45 degrees C, i.e. relevant temperatures for the clinical treatment of cancer. Heat shock-induced protein denaturation results in the aggregation of proteins to the nuclear matrix. Protein aggregation with the nuclear matrix is associated with the disruption of many nuclear matrix-dependent functions (e.g. DNA replication, DNA transcription, hnRNA processing, DNA repair, etc.) and cell death. Heat shock proteins are believed to bind denatured proteins and either prevents aggregation or render aggregates more readily dissociable. While many studies suggest a role for Hsp70 in heat resistance, we have recently found that nuclear localization/delocalization of Hsp70 and its rate of synthesis, but not its amount, correlate with a tumor cell's ability to proliferate at 41.1 degrees C. These results imply that not only is the nuclear matrix a target for the lethal effects of heat, but it also is a target for the protective, chaperoning and/or enhanced recovery effects of heat shock proteins., (Copyright Harcourt Brace and Co. Ltd 1998)

The present study was done to confirm the reported observation that low-intensity acute exposure to 2450 MHz radiation causes DNA single-strand breaks (Lai and Singh, Bioelectromagnetics 16, 207-210, 1995). Male Sprague-Dawley rats weighing approximately 250 g were irradiated with 2450 MHz continuous-wave (CW) microwaves for 2 h at a specific absorption rate of 1.2 W/kg in a cylindrical waveguide system (Guy et al., Radio Sci. 14, 63-74, 1979). There was no associated rise in the core body temperature of the rats. After the irradiation or sham treatments, rats were euthanized by either CO2 asphyxia or decapitation by guillotine (eight pairs of animals per euthanasia group). After euthanasia the brains were removed and immediately immersed in cold Ames medium and the cells of the cerebral cortex and the hippocampus were dissociated separately and subjected to the alkaline comet assay. Irrespective of whether the rats were euthanized by CO2 asphyxia or decapitated by guillotine, no significant differences were observed between either the comet length or the normalized comet moment of cells from either the cerebral cortex or the hippocampus of sham-treated rats and those from the irradiated rats. However, the data for the rats asphyxiated with CO2 showed more intrinsic DNA damage and more experiment-to-experiment variation than did the data for rats euthanized by guillotine. Therefore, the guillotine method of euthanasia is the most appropriate in studies relating to DNA damage. Furthermore, we did not confirm the observation that DNA damage is produced in cells of the rat cerebral cortex or the hippocampus after a 2-h exposure to 2450 MHz CW microwaves or at 4 h after the exposure.

The alkaline comet assay as described by Olive et al. (Exp. Cell Res. 198, 259-267, 1992) was used to detect DNA damage in cells exposed to low doses (0-5 cGy) of gamma radiation. Experiments were performed using lymphocytes isolated from whole blood of rats. The comet parameters, normalized comet moment and comet length, described by Kent et al. (Int. J. Radiat. Biol. 67, 655-660, 1995), were used as measurements of DNA damage. It was observed that the alkaline comet assay can detect DNA damage at doses as low as 0.6 cGy. The results of the experiments using low-dose gamma radiation are comparable with published results obtained using the alkaline comet assay according to the method of Singh et al. (Int. J. Radiat. Biol. 66, 23-28, 1994). Based on this observation and analysis of results published previously, we conclude that the version of the alkaline comet assay described by Olive et al. is as sensitive as other modifications of the comet assay reported in literature for the detection of DNA damage in cells exposed to low doses of ionizing radiation.

Recent reports suggest that exposure to 2450 MHz electromagnetic radiation causes DNA single-strand breaks (SSBs) and double-strand breaks (DSBs) in cells of rat brain irradiated in vivo (Lai and Singh, Bioelectromagnetics 16, 207-210, 1995; Int. J. Radiat. Biol. 69, 513-521, 1996). Therefore, we endeavored to determine if exposure of cultured mammalian cells in vitro to 2450 MHz radiation causes DNA damage. The alkaline comet assay (single-cell gel electrophoresis), which is reportedly the most sensitive method to assay DNA damage in individual cells, was used to measure DNA damage after in vitro 2450 MHz irradiation. Exponentially growing U87MG and C3H 10T1/2 cells were exposed to 2450 MHz continuous-wave (CW) radiation in specially designed radial transmission lines (RTLs) that provided relatively uniform microwave exposure. Specific absorption rates (SARs) were calculated to be 0.7 and 1.9 W/kg. Temperatures in the RTLs were measured in real time and were maintained at 37 +/- 0.3 degrees C. Every experiment included sham exposure(s) in an RTL. Cells were irradiated for 2 h, 2 h followed by a 4-h incubation at 37 degrees C in an incubator, 4 h and 24 h. After these treatments samples were subjected to the alkaline comet assay as described by Olive et al. (Exp. Cell Res. 198, 259-267, 1992). Images of comets were digitized and analyzed using a PC-based image analysis system, and the "normalized comet moment" and "comet length" were determined. No significant differences were observed between the test group and the controls after exposure to 2450 MHz CW irradiation. Thus 2450 MHz irradiation does not appear to cause DNA damage in cultured mammalian cells under these exposure conditions as measured by this assay.

Mouse C3H 10T1/2 fibroblasts and human glioblastoma U87MG cells were exposed to cellular phone communication frequency radiations to investigate whether such exposure produces DNA damage in in vitro cultures. Two types of frequency modulations were studied: frequency-modulated continuous-wave (FMCW), with a carrier frequency of 835.62 MHz, and code-division multiple-access (CDMA) centered on 847.74 MHz. Exponentially growing (U87MG and C3H 10T1/2 cells) and plateau-phase (C3H 10T1/2 cells) cultures were exposed to either FMCW or CDMA radiation for varying periods up to 24 h in specially designed radial transmission lines (RTLs) that provided relatively uniform exposure with a specific absorption rate (SAR) of 0.6 W/kg. Temperatures in the RTLs were monitored continuously and maintained at 37 +/- 0.3 degrees C. Sham exposure of cultures in an RTL (negative control) and 137Cs gamma-irradiated samples (positive control) were included with every experiment. The alkaline comet assay as described by Olive et al. (Exp. Cell Res. 198, 259-269, 1992) was used to measure DNA damage. No significant differences were observed between the test group exposed to FMCW or CDMA radiation and the sham-treated negative controls. Our results indicate that exposure of cultured mammalian cells to cellular phone communication frequencies under these conditions at an SAR of 0.6 W/kg does not cause DNA damage as measured by the alkaline comet assay.

The mode and kinetics of cell killing in HeLa and L5178Y cells were investigated to elucidate possible relationships between the mechanisms and modes of heat-induced cell death. L5178Y cells were heat-shocked for either 24 min at 43 degrees C or 30 min at 45 degrees C, while HeLa cells were given only the latter treatment. The degree of heat-induced excess nuclear protein correlated with cell death in HeLa cells but not in L5178Y cells. This difference suggests that the mechanism of cell death differs between these cell lines. With L5178Y cells the main mode of death after treatment at 43 degrees C was apoptosis with little progression of cells through the cell cycle. However, after treatment at 45 degrees C these cells died by necrosis without progressing through the cell cycle. HeLa cells were found to die by a mechanism other than apoptosis after significant progression of cells through the cell cycle and perturbation of the normal distribution of cells in the phases of the cell cycle (specifically, the fraction of cells in S and G2 phase increased 50% and 30%, respectively, prior to the occurrence of measurable cell death). These results are consistent with the hypothesis that the response to injury which has the potential to be lethal varies between different cell types, and results in the induction of different pathways leading to cell death.

Purpose: Transfection with either H-ras or H-ras and c-myc has been shown to confer radioresistance in rat embryonal cells (REC). REC primary, transfected with either c-myc, H-ras or cotransfected with c-myc and H-ras (in ascending order of radioresistance and tumorigenicity), were used as an in vitro model system to determine if nuclear matrix-mediated higher order DNA organization contributes to oncogene-mediated radioresistance., Methods and Materials: DNA damage induction and repair were measured by the alkaline and neutral filter elution assays. Analysis of the ability of DNA loop domains to undergo supercoiling changes in the presence of radiation-induced damage was determined by the fluorescent halo assay (FHA). Because DNA loops are organized by the nuclear matrix (NM), a study of NM-associated proteins by high resolution two-dimensional gel electrophoresis was performed., Results: Induction and repair rates of DNA single- and double-strand breaks were similar for the relatively radiosensitive c-myc transfected and the radioresistant c-myc + H-ras transfected cells. However, the degree of inhibition of DNA supercoil rewinding in the presence of radiation-induced damage was less in the radioresistant cells and was inversely correlated with survival. A progressive loss of NM-associated proteins was observed, which correlated with increasing radioresistance and tumorigenicity in these cell lines. In addition, some protein changes were consistent with the possibility that these changes could be involved in DNA anchoring., Conclusions: Increased radioresistance associated with increasing tumorigencity in these oncogene-transfected cell lines could be due to changes in NM-mediated DNA organization, possibly via differences in NM protein composition that occur following oncogenic transfection.

Cell of the radioresistant L5178Y-R, -S35, -SR and M10(neo 5)-1 and radiosensitive L5178Y-S, M10 and LX830 cell lines were used to investigate the relationship between radiosensitivity and DNA supercoiling ability mediated by the nuclear matrix within chromatin loops containing DNA damage. The ability of DNA loops to undergo changes in supercoiling in the presence of radiation-induced damage revealed that in all cases the degree of inhibition of supercoil rewinding was greater in the radiosensitive cells. Since the amount of DNA damage induced per unit dose is known to be equal in all these cell lines, the same number of DNA lesions produced a greater loss of topological constraint in the radiosensitive cells. The differential loss of DNA supercoiling ability could be due to differences in DNA-nuclear matrix anchor points. High-resolution two-dimensional gel electrophoresis of nucleoid proteins showed numerous reproducible differences in nuclear matrix protein between the cell lines studied. A total of nine proteins were associated with nucleoids from L5178Y-R cells and absent from L5178Y-S nucleoids. None of them, however, correlated absolutely with radioresistance. Thus, unlike previous studies in CHO cells, no candidates for the conveyance of cellular radiosensitivity that were single proteins were detected. However, these results are consistent with the hypothesis that stability of DNA loop domains in the presence of DNA damage is a determinant of the outcome of radiation-induced DNA damage.

We have investigated the composition of nuclear matrix proteins and DNA supercoiling characteristics of cell lines expressing altered radiation sensitivity. Chinese hamster ovary cell lines 4364 (wild-type), XR-1 (DSB repair-deficient, radiosensitive) and XR-122 (a radioresistant variant of XR-1 bearing human chromosome 5) were used as a model to study the relationship between intrinsic radiation sensitivity and the level of DNA supercoiling ability within chromatin loops and the composition of nuclear matrix proteins. Analysis of the ability of DNA loop domains to undergo changes in DNA supercoiling in the presence of DNA damage revealed that the degree of inhibition of loop rewinding was greater in the radiation-sensitive cells (XR-1) compared to the radiation-resistant cells (4364 and XR-122). Furthermore, the loop-rewinding characteristics correlated inversely with the clonogenic survival of these cells after exposure to ionizing radiation. Since DNA loops are anchored to the nuclear matrix by protein-DNA anchor points, a study of the nuclear matrix proteins by high-resolution 2D-PAGE was conducted for these cells to determine whether differential inhibition of loop rewinding could be due to differences in the DNA loop-protein anchor points in these cells. The XR-1 cells showed an overall absence of 13 proteins compared to the 4364 cells. Of these 13, 5 were restored in XR-122 cells. These results are consistent with the hypothesis that stability of the DNA loop domains in the presence of DNA damage contributes to the expression of potentially lethal damage by ionizing radiation.

The cell-cycle age response to 44 or 45 degrees C hyperthermic treatment was studied in M10, a mutant of mouse L5178Y cells, and human Burkitt P3HR-1 cells synchronized by centrifugal elutriation. Survival response to 44 degrees C hyperthermia or radiation showed that Burkitt cells were relatively hyperthermic resistant and average in radiosensitivity, while M10 cells were relatively hyperthermic sensitive and substantially radiosensitive. A typical age-response to hyperthermia through the cell-cycle was observed in M10 cells, although no significant variation in the response to hyperthermia was demonstrated in Burkitt P3HR-1 cells. Combined effect of procaine with 44 degrees C 20-min treatment for Burkitt P3HR-1 cells showed a substantial enhancement of cytotoxicity in S phase cells. The probable implication of membrane composition, membrane changes due to heat and its modifying agents are discussed. Furthermore, when hyperthermia was combined with radiation in Burkitt P3HR-1 cells, cytotoxic enhancement was observed in G1/S boundary phase, not in S phase.

Cell lysis and eosin staining were observed in L5178Y cells within the first 3 h of post-hyperthermia incubation at 37 degrees C, after which both leveled to a plateau. Lysis and eosin staining were proportional to the severity of heat in asynchronous cells, whereas it was maximum in the most heat-sensitive M phase, intermediate in S, and least in heat-resistant G1 for the same heat treatment. Further, leakage of labeled [3H]thymidine and a decrease in radioactivity retained within heated cells coincided with an increase in eosin staining, indicating that the dye uptake was due to membrane damage. It was presumed that the eosin-stained fraction represented dead cells. The percentage eosin-stained cells reached a plateau, and this level was used to determine survival; when the results were compared with those obtained by the colony formation method, they were identical. By comparing the two survival assay methods we concluded that cell death after hyperthermia in L5178Y cells is mainly by interphase death in all phases of the cell cycle. The reasons for this conclusion are that a reduction in survival could be detected within one generation of L5178Y cells by the eosin staining method, and the survival values obtained by this method were identical to those obtained by the colony formation method.