Robert S. Malyapa

DNA Damage in Rat Brain Cells after In Vivo Exposure to 2450 MHz Electromagnetic Radiation and Various Methods of Euthanasia

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.

Nuclear matrix as a target for hyperthermic killing of cancer cells

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 cells 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.

Skin cancer of the head and neck with perineural invasion.

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.

Definitive Radiotherapy for Tonsillar Squamous Cell Carcinoma

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.

Detection of DNA damage by the alkaline comet assay after exposure to low-dose gamma radiation

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.

Comparison of Molecular Markers of Hypoxia and Imaging with 60 Cu-ATSM in Cancer of the Uterine Cervix

PurposeTo determine if hypoxia-related molecular markers are associated with 60Cu labeled diacetyl-bis (N4-methylthiosemicarbazone); (60Cu-ATSM) imaging of tumor hypoxia in cervical cancer.ProceduresFifteen patients were enrolled in a prospective study and underwent evaluation of tumor hypoxia with positron emission tomography (PET) using 60Cu-ATSM. 60Cu-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.ResultsSix patients had hypoxic tumors determined by 60Cu-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 60Cu-ATSM to be a significant independent predictor of tumor recurrence (p = 0.0287).Conclusions60Cu-ATSM hypoxia was correlated with overexpression of VEGF, EGFR, COX-2, CA-9, an increase in apoptosis, and a poor outcome.

Salivary Gland Pleomorphic Adenoma

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.

Adjuvant interferon-based chemoradiation followed by gemcitabine for resected pancreatic adenocarcinoma: a single-institution phase II study.

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-α (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-α with 5-fluorouracil-based chemoradiation. Despite these encouraging results, significant concerns regarding dose- and treatment-limiting toxicities remain.

Thymidylate Synthase Genotype-Directed Neoadjuvant Chemoradiation for Patients With Rectal Adenocarcinoma

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.

Carcinoma of the nasal cavity and paranasal sinuses

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.