Effect of whole-body hyperthermia and cyclophosphamide on natural killer cell activity in murine erythroleukemia.

Mice infected with the polycythemia-inducing strain of Friend virus complex (FVC-P) develop a fatal erythroid disease similar in some respects to leukemia. Six- to eight-week-old DBA/2 female mice were injected i.v. with 0.5 ml of a virus suspension containing approximately 5 X 10(4) plaque-forming units and 5 X 10(3) spleen focus-forming units. Four treatment regimens were begun 3 days postinjection: (a) no treatment; (b) whole-body hyperthermia (WBH) alone; (c) cyclophosphamide (CY) alone; (d) WBH combined with CY. WBH treatment utilized a microwave generator operating at 2450 MHz. The i.p. temperature of the mice receiving WBH was maintained at 39.5-40 degrees C for 30 min. The CY was given i.p. at a dosage of 20 mg/kg of body weight. The various treatments, CY, WBH, CY + WBH were given once a week for 2 weeks. Natural killer cell activity was examined in all four groups of mice and was found to be significantly higher in the animals treated with WBH or CY. Our results show that WBH, either alone or in combination with CY, can prolong the lifespan of mice infected with lethal dosages of the FVC-P, possibly via a mechanism involving natural killer cells.

Effects of interleukin 2 treatment combined with local hyperthermia in mice inoculated with Lewis lung carcinoma cells.

Recombinant human (rhu) interleukin 2 (IL-2) was evaluated alone and in combination with local hyperthermia (LH) in mice inoculated s.c. with 5 x 10(5) Lewis lung carcinoma cells. Four treatment regimens were begun 6 days postinoculation at a time when the tumor had grown to approximately 8.0 mm in diameter. Treatments were: group 1, saline injected as control; group 2, LH; group 3, rhuIL-2; or group 4, LH combined with rhuIL-2. LH utilized hot water circulation by a Brann Thermomix 1420. The intratumor temperature was maintained at 43 +/- 0.2 degrees C for 30 min each on days 6 and 10 and rhuIL-2 was given s.c. at 5 x 10(4) units twice a day for 5 days. Thirty mice in each group were sacrificed 28 days after tumor inoculation. An additional 20 mice in each group were observed for survival time. The size of primary tumor and the number of lung metastases were reduced and the survival time was prolonged in mice treated by either LH or IL-2. However, a greater antitumor effect in Lewis lung carcinoma tumor-bearing mice was observed using IL-2 therapy combined with LH. Tumor growth was associated with increased splenic granulocyte-macrophage progenitor cells and an abnormal L3T4+/Lyt-2+ lymphocyte subset ratio (less than 1.0). Splenic granulocyte-macrophage progenitor cell numbers and the L3T4+/Lyt-2+ ratio returned to normal in the group treated with combination therapy, the best responder group. The L3T4+/Lyt-2+ ratio did not change in the groups treated with single therapy. These results suggest the efficacy and possible clinical relevance of combined therapy with rhuIL-2 and LH for certain metastatic tumors.

Influence of elevated temperature on natural killer cell activity, lymphokine-activated killer cell activity and lectin-dependent cytotoxicity of human umbilical cord blood and adult blood cells.

PURPOSE: To determine whether hyperthermia is to the benefit or detriment of host immune function, the effect of hyperthermia was evaluated on various functions of T-lymphocytes from human umbilical cord blood and compared to that of adult blood. METHODS AND MATERIALS: Nonadherent mononuclear cells from cord blood or adult blood were used as the effector cells. To generate lymphokine activated killer (LAK) cells, effector cells were kept in culture for 5 days in complete medium containing recombinant human interleukin-2. To activate effector cells to become cytotoxic, cells were kept in culture in complete medium containing Con A. Cytotoxicity was determined in a standard 4-h chromium release assay using K-562 human erythroleukemic cells (in the natural killer cell activity assay) or Daudi cells (in the LAK cell activity or Lectin dependent cytotoxicity assay) as targets. For heat effects, cells in complete medium were heated at the desired temperature in a water bath for 1 h. RESULTS: Lymphokine-activated killer cell activity, lectin-dependent cytotoxicity and T-cell proliferative capacity were not deficient in human cord blood. Cytotoxic activities of T-cells from adult blood as well as from cord blood can be enhanced at febrile range (< or = 40 degrees C), and were significantly decreased by exposure to 1 h at 42 degrees C. CONCLUSION: The febrile responses (< or = 40 degrees C) to infection, in the course of malignant disease and with biological response modifiers treatment, may all be related to host defense mechanisms. Based on these observations, whole body hyperthermia (< or = 40 degrees C), in combination with the appropriate cytokines, may have therapeutic potential in the treatment of neonatal infections and malignancies under certain circumstances. Hyperthermia in febrile range may, therefore, confer an important immunoregulatory advantage to the host. In contrast, tumor killing therapeutic temperature (> 42 degrees C) which inhibits host immunocompetence should probably be used only for local hyperthermia.

Whole body hyperthermia: a potent radioprotector in vivo.

Interleukin-1 has been reported to be an effective radioprotective agent in mice subjected to lethal doses of irradiation. Production of Interleukin-1 can be increased by whole body hyperthermia. Therefore, whole body hyperthermia was assessed for its efficacy in protecting the lethal effects of ionizing radiation in DBA/2 mice. One hour of 40 degrees C +/- 0.2 whole body hyperthermia given 20 hr before 900 cGy total body irradiation protected 100% of DBA/2 mice from an LD 100/16 radiation dose (dose of irradiation that killed 100% of the mice in 16 days). Lethal doses of total body irradiation produced profound monocytopenia, decreased cellularity of thymus, spleen, and bone marrow, and suppressed Interleukin-1 production. Interleukin-1 production was determined using the thymocyte proliferation assay. Whole body hyperthermia accelerated recovery of blood leukocytes by up to 5 days post-total body irradiation in DBA/2 mice. Thymocytes, spleen, and bone marrow cells were activated by whole body hyperthermia, as assessed by the cell's response to Concanavalin A. This was accompanied by accelerated Interleukin-1 generation. Our results provide the first evidence that whole body hyperthermia acts as a potent radioprotector in vivo, effects that may be mediated by Interleukin-1.

A reliable method for quantitating chromatin fragments by flow cytometry to predict the effect of total body irradiation and hyperthermia on mice.

The frequencies of chromatin fragments, including micronuclei, in murine thymus cells, spleen cells and bone marrow cells have been used as a quantitative indicator of gamma-ray induced chromosome damage and could be used to screen potential radioprotective agents as well. The yield of chromatin fragments induced in mice receiving different dosage levels of total body irradiation alone and in mice also given whole body hyperthermia as a potent radioprotector were assessed by flow cytometric analysis. Our results demonstrated that chromatin fragments induced by irradiation in vivo was clearly dose-dependent and that chromatin fragments could potentially serve as a biological indicator of radiation damage. One hour of whole body hyperthermia at 40 degrees C (+/- 0.2 degree C) given 20 hours before a lethal dosage (900 cGy) of total body irradiation protects 100% of DBA/2 mice from an LD 100/16 irradiation dose (dose of irradiation that killed 100% of the mice in 16 days). This is in good agreement with the percent of chromatin fragments formed in the cells of the protected animals, which showed no significant difference from those observed in the normal mice. The results indicate that whole body hyperthermia protected the thymus and bone marrow from irradiation damage. This study provides further evidence which supports that whole body hyperthermia can act as a potent radioprotector in vivo. Measurement of the frequencies of chromatin fragments by flow cytometry is simple and reliable. The method can be applied to screen radioprotective agents.

A comparison of lung metastases and natural killer cell activity in daily fractions and weekly fractions of radiation therapy on murine B16a melanoma.

C57BL/6J male mice were inoculated with 5 X 10(5) B16a melanoma cells. Seven days post-inoculation, when the tumor had grown to 8.0-10.0 mm in diameter, 120 tumor-bearing mice were randomly divided into three groups: (1) sham-irradiated controls, (2) mice receiving 200 cGy five times a week for 6 weeks, and (3) mice receiving 800 cGy once a week for 4 weeks. Thirty mice in each group were sacrificed 47 days postinoculation. Ten mice in each group were observed for the survival time data. The primary tumor was significantly smaller and the number of lung metastases were significantly fewer in mice treated with 800 cGy once a week compared to mice treated with 200 cGy five times a week. When natural killer (NK) cell activity was assessed against YAC-1 tumor targets, it was found to be significantly higher in mice treated with a single large weekly dose of irradiation. These results show that B16a melanoma responds more favorably to a single large dose of irradiation administered once a week compared to the smaller conventional fraction administered five times a week. This beneficial effect correlates with an increase in NK activity, indicating that there may be a causal relationship.

Editorial: Post-therapeutic radiation injuries of the nervous system. Reflections on their prevention.

The introduction of the concept of Nominal Standard Dose and of Time, Dose Fractionation Factors ostensibly permits definition of tolerance doses for normal tissues in unequivocal terms. However, even with these refinements, tolerance doses remain, at best, guidelines, because radiobiologic effectiveness is governed not only by the effective dose, but also by individual factors, which will modify the response. Attention must be accorded to these biologic parameters, in order to prevent injury to healthy tissues. Of particular significance are the relative size or volume of the irradiated tissue, the possible presence of co-existing pathology in the exposed organ and the development of disease after tge termination of the treatment. Even if these factors are properly respected, the risk of radiation injury cannot entirely be eliminated. The radiotherapist is therefore obligated to use an approach which minimizes the exposure of the healthy nervous tissue, a goal which has become attainable with the advent of modern accelerators as radiation sources.

Radiotherapy in the treatment of verrucous carcinoma of the head and neck.

Because of reports of anaplastic transformation following irradiation, this study examines the incidence of anaplastic transformation and local control of these lesions. This review of seven patients who had verrucous carcinoma of the head and neck that was treated with irradiation shows local control in 71% of cases. There were no cases of anaplastic transformation. This report adds to the literature two cases of "de-differentiation" to less differentiated squamous carcinomas; one such case occurred after surgery alone. The literature is reviewed. Overall, anaplastic transformation is reported in 7% of patients who had irradiation. De-differentiation occurs after surgery as well. The rate of local control with irradiation is less than 50%; with surgery it is 85%. It is concluded that surgery should be used if the procedure has acceptable morbidity. Otherwise, irradiation can be used. Failures can be salvaged surgically. "Anaplastic transformation" should not affect treatment approach.

Epithelial carcinoma of the nasal fossa.

Epithelial carcinoma of the nasal cavity and vestibule occurs relatively rarely and is primarily treated with surgical excision, radiation therapy, or a combination of these two treatment modalities. This review describes the treatment methods and results of patients seen at the Indiana University School of Medicine from 1963 through 1983 in the Departments of Radiation Oncology and Otolaryngology. Using the medical records of 34 patients, this retrospective study reviews the clinical significance of histology, anatomical location of primary tumors, and therapeutic results, including the pattern of treatment failures of primary tumors. The results of radiation therapy alone compare favorably with the results of surgery alone. A review of the literature and this study indicate that patients receiving a combination of surgery and irradiation face a better prognosis than those who experience surgery or irradiation alone.

The role of radiation therapy in the treatment of malignant tumors of the paranasal sinuses.

During a 20 year period, 121 patients with malignant tumors of the paranasal sinuses were seen at this institution. Of these cases, 109 patients with carcinomas originating from the mucous or paranasal sinuses were evaluated. Data were analyzed on the basis of 1. primary tumor control and metastatic nodal sterilization, 2. pre and post-therapy incidence and location of neck node metastasis, and 3. overall survival. Data were also analyzed as to survival on the basis of TNM classification and histology. From this retrospective study, the incidence of neck node metastasis prior to treatment appears to be slightly higher than in literature (23/88 or 26%). The incidence of post-treatment metastasis is 9/88 or 10%. No significant correlation was noted between histology and neck node metastasis.

Wound complications associated with brachytherapy for primary or salvage treatment of head and neck cancer.

Brachytherapy can be employed in the primary or salvage treatment of head and neck cancer. The advantage of brachytherapy is the stereotactic limitation of radiation exposure to noninvolved tissues. Wound complications associated with brachytherapy have been discussed only sporadically in the literature. This retrospective study examines 28 patients, 20 for initial treatment and eight for salvage, with varying site and stage head and neck cancer treated with brachytherapy in addition to external beam radiation therapy and/or surgery. The overall complication rate was 50% (14/28), with infection and minor flap breakdown being the most common problems. Tumor site in the primary treatment group was the only significant factor in wound complications. In the salvage group complications were minor and primarily related to flap coverage of brachytherapy catheters.

Comparison of radiation therapy of classic and epidemic Kaposi's sarcoma.

Between 1963 and 1990, 92 lesions in twenty patients with Kaposi's sarcoma (KS) were treated using radiation therapy (RT). Fifty-nine classic Kaposi's sarcoma (CKS) lesions in 12 patients were treated with 990 cGy in 3 fractions-5,000 cGy in 25 fractions; 39 lesions (66%) showed a complete response (CR), 15 (25%) showed a partial response (PR) with complete symptomatic relief, and 5 lesions (8%) showed no response (NR). Only 6 of 59 lesions (10%) recurred in field with a median recurrence-free duration of 8 months. Thirty-three epidemic Kaposi's sarcoma (EKS) lesions in 8 patients were treated from 1,000 cGy in 5 fractions-3,000 cGy in 10 fractions. Thirty-one EKS lesions (94%) showed CR, and two lesions (6%) showed PR. An extensive review of the literature is also presented in the study here reported.

Extranodal lymphoma of the head and neck area.

Records of 64 patients with extranodal lymphoma of the head and neck area treated at this institution between 1965 and 1982 have been reviewed. Of these cases, 78% had Stage I-EA disease, 20% had Stage II-EA disease, and 2% had Stage III-EA disease. Excluded from this study were patients of the pediatric age, patients with previous chemotherapy and surgery, patients in whom extranodal involvement has been a terminal manifestation of wide-spread disease, and patients with nodal involvement with extension of disease outside the lymph nodes. Patients with a histiocytic histology accounted for 63%, 28% were of lymphocytic histologies, 4.5% were of mixed histologies, and in 4.5% a diagnosis of lymphoma or benign lymphocytic proliferation had been made. The paranasal sinuses, nasal cavity, and oral cavity were the most frequent sites of involvement, making up 39% of the cases--with the orbit, central nervous system, nasopharynx, salivary glands, thyroid, skin, and larynx following in that order of frequency. All patients received a definitive course of radiotherapy, except patients with CNS involvement. In no case was chemotherapy given concomitantly with the radiation therapy. One-third of the patients did require chemotherapy at a later time for progression of disease. No patients were lost to follow-up. Seven patients with primaries of the brain were inevaluable for response; however, all patients having visible or palpable tumors achieved a complete response (100%), and there was no recurrence or persistent disease in the field of therapy. The next area of disease involvement depended on the site of the primary. Nodal involvement following extranodal disease carried a poor prognosis.

Total skin electron beam radiation therapy for mycosis fungoides.

Forty-nine patients with biopsy-proven mycosis fungoides, Stages I-IV were treated using total skin electron beam irradiation (TSEBI). Total dose ranged from 600 cGy to 3,200 cGy. To evaluate the dose response relationship, patients were retrospectively divided into two groups. In Group I, 18 patients received a dose of 2,000 cGy or less, and in Group II, 31 patients received more than 2,000 cGy. The overall response rate was 87.7% with a 75.7% complete response and 12.2% partial response. Complete response was higher among patients with early stage disease: (Stage IA 1/1, Stage IB 23/35 (92%), Stage IIA 3/4 (75%), Stage IIB 4/8 (50%), Stage III 3/6 (50%), Stage IVA 1/1, Stage IVB 0/1, and unstaged group 2/3 (66.6%)). Patients treated with a higher total dose had a higher overall 5-year survival rate (Group I 38%, Group II 68%), longer median duration of complete response (Group I, 27 months; Group II, 35.3 months), slightly better complete response rate (72.2% for Group I, 77.4% for Group II), and lower recurrence rate (Group I, 94%; Group II, 83.9%) compared to patients with lower total dose. Complications from TSEBI were minimal. Total skin electron beam irradiation is effective in controlling early stage mycosis fungoides; however, a prospective study to evaluate optimum total dose is needed.

Synergistic cytotoxic and antitumor effects of irradiation and taxol on human HeLa cervix carcinoma and mouse B16 melanoma cells.

Several articles have appeared in the scientific literature which report that taxol has the ability to block and/or prolong cells in the G2/Mp phase of the cell cycle by inducing extremely stable microtubules. The G2/M phase is known to be the most radiosensitive phase of the cell cycle. It is the purpose of this study to evaluate the effect of combination taxol and ionizing radiation on tumor cell lines which have not been previously reported in the literature. Decrease in viability and inhibition of proliferation of HeLa and B16 cells induced by irradiation were dose-dependent and significantly enhanced by pretreatment the cells with taxol. Similar antitumor effects of irradiation and taxol were demonstrated by flow cytometric analysis of chromatic fragment formation induced in these cells. The exact mechanism by which taxol enhanced the tumoricidal effect of irradiation is not known. Cell cycle analysis showed that taxol was effective in blocking HeLa and B16 cells at the G2/M stage, at which the tumor cells are believed to be most sensitive to irradiation. This study is in agreement with others who have found that taxol is a powerful radiation sensitizer. Clinical research protocols have been developed and are under way to determine if taxol produces similar synergistic effects in patients undergoing radiation therapy.

An overview of the role of radiation therapy and hyperthermia in treatment of malignant melanoma.

From January, 1970 until December, 1987, a total of 188 malignant melanoma lesions in 92 patients were treated at the Department of Radiation Oncology, Indiana University Medical Center, Indianapolis, Indiana. Response was evaluated in 181 evaluable lesions treated by radiation alone and radiation plus hyperthermia to assess differences in response to a total dose, dose per fraction and overall time of treatment, as well as effects of adjunctive hyperthermia treatment. Different fractions of radiation, ranging from 100 cGy to 1000 cGy, were used. Local hyperthermia was administered for one hour following radiation treatment using microwave with different frequencies. The tumor temperature was also monitored during treatment. With a radiation dose of less than 400 cGy per fraction, and complete response rate (CR) was 34% (16/47) and the objective response rate (OR) was 62% (29/47). When hyperthermia was added, the complete response rate rose from 34% to 70%. With a dose of more than 400 cGy per fraction, the CR was 63% (48/77), and OR was 95% (73/77). When hyperthermia was added, the complete response rate rose from 63% to 77%.

Micronuclei assay--a predictive variable for tumor response to treatment.

Our preliminary data indicate that the formation of micronuclei (MN) in treated tumor cells is a predictive variable for tumor response to treatment. In a pilot study involving four patients who received both radiation therapy and hyperthermia, fine needle aspirate (FNA) samples were taken and analyzed before therapy, and after each 1000 centigray (cGy) up to 3000 cGy. The results indicate a correlation between increasing formation of micronuclei and decreasing tumor volume. All of the patients in this Study have had their tumors under control for at least one year. Our preliminary data demonstrated that a high level of micronuclei in tumor cells correlates with favorable response of the tumor to treatment with radiation and heat. The assay is easy to perform and FNA biopsy could be done in the clinic with minimal discomfort to the patient.

Rhabdomyosarcoma in the pediatric age group.

Forty-five consecutive cases of pediatric rhabdomyosarcoma were studied as to age at onset, histological type, primary site, stage of disease when first seen, method of treatment, and survival. It appears that rhabdomyosarcoma is a relatively radiosensitive tumor which can be controlled locally with radiation alone or in combination with surgery to excise the bulk of the tumor mass. On the other hand, chemotherapy must necessarily play a major role in treatment since most cases are first seen in the late stages of the disease. Close cooperation among a wide range of specialists, such as the pediatrician, radiation therapist, pediatric oncologist, pathologist, and surgeon is essential to accomplish a maximum therapeutic effect in the unfortunate children who develop this relatively rare but aggressive tumor.

Carcinoma of the lower lip: treatment results at Indiana University Hospitals.

Sixty-six consecutive cases of carcinoma of the lower lip were seen at the Indiana University Hospitals from 1960 to 1972. Each patient's records were reviewed as to age, sex, race, stage, stage by treatment modality, pathology, treatment methods used and cause of failure. Early lip cancer represents a highly curative lesion whether treated by surgery or radiation therapy. Close followup is mandatory on all patients as the development of neck nodes, if discovered late, carries a grave prognosis. Advanced carcinoma of the lip can be successfully treated with current radiation therapy techniques using fractionated doses providing excellent cosmetic results.