HDAC1,2 inhibition and doxorubicin impair Mre11-dependent DNA repair and DISC to override BCR-ABL1-driven DSB repair in Philadelphia chromosome-positive B-cell precursor acute lymphoblastic leukemia.

Philadelphia chromosome-positive (Ph+) B-cell precursor acute lymphoblastic leukemia (ALL) expressing BCR-ABL1 oncoprotein is a major subclass of ALL with poor prognosis. BCR-ABL1-expressing leukemic cells are highly dependent on double-strand break (DSB) repair signals for their survival. Here we report that a first-in-class HDAC1,2 selective inhibitor and doxorubicin (a hyper-CVAD chemotherapy regimen component) impair DSB repair networks in Ph+ B-cell precursor ALL cells using common as well as distinct mechanisms. The HDAC1,2 inhibitor but not doxorubicin alters nucleosomal occupancy to impact chromatin structure, as revealed by MNase-Seq. Quantitative mass spectrometry of the chromatin proteome along with functional assays showed that the HDAC1,2 inhibitor and doxorubicin either alone or in combination impair the central hub of DNA repair, the Mre11-Rad51-DNA ligase 1 axis, involved in BCR-ABL1-specific DSB repair signaling in Ph+ B-cell precursor ALL cells. HDAC1,2 inhibitor and doxorubicin interfere with DISC (DNA damage-induced transcriptional silencing in cis)) or transcriptional silencing program in cis around DSB sites via chromatin remodeler-dependent and -independent mechanisms, respectively, to further impair DSB repair. HDAC1,2 inhibitor either alone or when combined with doxorubicin decreases leukemia burden in vivo in refractory Ph+ B-cell precursor ALL patient-derived xenograft mouse models. Overall, our novel mechanistic and preclinical studies together demonstrate that HDAC1,2 selective inhibition can overcome DSB repair 'addiction' and provide an effective therapeutic option for Ph+ B-cell precursor ALL.

Effect of glutathione depletion by L-buthionine sulfoximine on the cytotoxicity of cyclophosphamide in single and fractionated doses to EMT6/SF mouse tumors and bone marrow.

The combined cytotoxicity of cyclophosphamide [(CYC) CAS: 50-18-0] and glutathione [(GSH) CAS: 70-18-8] depletion by buthionine sulfoximine (BSO) toward EMT6/SF tumors and mouse bone marrow was studied in male BALB/c mice. Tumor GSH was depleted to 28.7 and 7.8% of control by 1 or 2 doses of BSO (5 mmol/kg), respectively, administered ip at 12-hour intervals prior to assay. Tumor GSH could be maintained at a level 8% of control by daily dosing with BSO for 3 days. The same BSO administration schedule lowered bone marrow cell GSH to 31% of control for the 3-day period studied. Tumor growth and bone marrow cell counts were unaffected by all BSO treatments studied. However, BSO pretreatment enhanced the cytotoxicity of CYC to tumor cells, as measured by an in vitro colony-forming assay, but it did not enhance the depletion of bone marrow cells in CYC-treated mice. The magnitude of the enhancement of CYC cytotoxicity by BSO depended on the extent of GSH depletion, tumor size, and the CYC dosing schedule: Single and double doses of BSO enhanced the cytotoxicity of a single dose of CYC by factors of 1.5 and 2.0, respectively, in 7-day-old tumors, based on the dose of CYC required to produce a surviving fraction of 4 X 10(-3). The response of 9-day-old tumors to CYC suggested the presence of a subpopulation of cells that were relatively resistant to CYC. There was no evidence for this subpopulation in BSO-pretreated tumors. Multiple doses of BSO and CYC also combined to give enhanced (1.65-fold) tumor cell killing compared to tumors treated with CYC alone. The data suggest a possible role for BSO as a clinical chemosensitizer, which could be combined with fractionated doses of CYC and may be effective on small cancerous lesions.

Effects of extreme hypoxia on the growth and viability of EMT6/SF mouse tumor cells in vitro.

The purpose of this study was to characterize a model system in which to study hypoxic cell biology in vitro as a function of time under extremely hypoxic conditions. EMT6/SF cells that were maintained at 37 degrees under hypoxic conditions showed no increase in cell number for up to 70 hr. The mitotic index of hypoxic cultures was less than 0.1%, compared to 2.3 to 3.0% in aerated cultures. The plating efficiency of hypoxic cells decreased with time to 20 to 30% of control values by 70 hr. Aerated cultures consumed glucose more rapidly than did hypoxic ones, due to increasing cell number in air. But, on a per cell basis, hypoxic and aerated cells consumed glucose at equal rates (congruent to 1.2 X 10(-4) micrograms/cell/hr). Virtually 100% of the glucose consumed was converted into lactic acid in both aerated and hypoxic cultures. The labeling index and rate of incorporation of [3H]thymidine decreased exponentially with time in hypoxia. However, the percentage of cells with S-phase DNA content remained nearly constant for up to 72 hr. The rate of protein synthesis was suppressed in hypoxic cultures to between 20 and 50% of control (aerated) rates. When cultures were reaerated following 45 hr of hypoxia, congruent to 12 hr was required for resumption of DNA synthesis and cell division. The application of this system to further study of hypoxic cell biology is discussed.

Effects of hypoxia, pH, and growth stage on cell killing in Chinese hamster V79 cells in vitro by activated cyclophosphamide.

Several factors which influence the sensitivity of Chinese hamster V79 cells to cyclophosphamide (CY) have been studied in vitro in both suspension and monolayer cultures. Activated CY was obtained from the blood of mice 15 to 30 min after i.p. injection of CY (400 mg/kg). At pH 7.4, hypoxia rendered the cells more sensitive to activated CY. At lower values of pH (6.6 and 7.0), there was no difference between the sensitivities of oxic and hypoxic cells, although cells in both conditions were more sensitive to CY than at pH 7.4. Drug sensitivity was markedly affected by the stage of cell growth. Monolayer cultures were most sensitive to CY within a few hours of plating. Cultures then rapidly became less sensitive, with maximum resistance occurring between 24 and 48 h after plating, while the cells were still exhibiting rapid exponential growth. This development of resistance parallelled the formation of small colonies (2 to 4 cells), implying that intercellular contact may confer resistance to killing by activated CY.

Cellular glutathione, thermal sensitivity, and thermotolerance in Chinese hamster fibroblasts and their heat-resistant variants.

HA-1 Chinese hamster fibroblasts and two heat-resistant variants, designated 2242 and 3012. have been investigated to determine the role that glutathione (GSH) plays in intrinsic cellular resistance to heat and in the development of thermotolerance. The constitutive levels of GSH did not correlate with intrinsic heat sensitivities, but depletion of GSH sensitized all three cell lines to thermal stress. After heating (43.5 degrees C/2 h), surviving fractions were 1 X 10(-3), 1 X 10(-2), and 8 X 10(-3) for HA-1, 2242, and 3012 cells, respectively. Depletion of cellular GSH with L-buthionine-S, R-sulfoximine to less than 10% of control values sensitized such that the thermal responses of these three cell lines were nearly indistinguishable at 43.5 degrees C. Surviving fractions were 2 X 10(-4), 1 X 10(-4), and 1 X 10(-4) for L-buthionine-S,R-sulfoximine-treated HA-1, 2242, and 3012 cells, respectively, following heating at 43.5 degrees C for 2 h. The development of thermotolerance in HA-1 cells following heat shock (45 degrees C/15 min) was unaffected by the inhibition of GSH synthesis. On the other hand, when GSH levels were maintained at extremely low levels, the development of thermotolerance was inhibited. In addition, following heat shock, cellular GSH was decreased and remained below control levels during the development of thermotolerance.

A novel alternative approach for prediction of radiation response of squamous cell carcinoma of head and neck.

Accurate prediction of human tumor response to radiation therapy and concomitant chemoradiation would be an important tool to assist the physician in making recommendations for tumor treatment. Most of the studies that define the molecular markers for prediction of radiation response are based on the observation of gene expression using immunostaining, Northern blot, or Western blot analysis of a single or several genes. The results vary among different studies, and some results are contradictory. However, the studies agree that the change in expression of the tumor-related gene affects the radiation response. In this study, we explored a novel approach to predict the radiation response of human tumor using Atlas human cancer 1.2 cDNA array to analyze the expression profile of 1187 tumor-related genes in radiation-resistant and radiation-sensitive tissues. Sixty tumor-related genes were selected as predictors of radiation response of squamous cell carcinoma of the head and neck. Using the expression intensity of these 60 tumor-related genes, in combination with cluster analysis, we successfully predicted the radiation identity of two tumor samples.

Quantitative analysis of cellular glutathione by flow cytometry utilizing monochlorobimane: some applications to radiation and drug resistance in vitro and in vivo.

An assay using a bimane derivative has been developed to detect free glutathione (GSH) in individual viable cells by flow cytometry. Monochlorobimane [syn-(ClCH2CH3)-1,5-diazabicycla[3.30]acta-3,6-diene-2,8-dio ne], itself nonfluorescent, reacts with GSH to form a highly fluorescent derivative. High pressure liquid chromatography analysis showed that, using specific staining conditions, the only low molecular weight fluorescent derivative formed in Chinese hamster ovary cells was that formed with GSH. Very little reaction with protein sulfhydryls was observed. Rates of GSH depletion in Chinese hamster ovary cells exposed to diethylmaleate were essentially the same, whether measured by relative fluorescence intensity, by flow cytometry or by enzymatic assay on cellular extracts. This method was shown to be useful for measurement of GSH resynthesis, uptake, and depletion by prolonged hypoxia and misonidazole treatment. Since measurements are made on individual cells, cell-to-cell variation and populational heterogeneity in GSH content are revealed by flow cytometry. Although under most conditions in vitro GSH content is relatively homogeneous, under certain circumstances, such as release from hypoxia, heterogeneity in populational GSH levels was observed. The significance of this heterogeneity is discussed in regard to the induction of gene amplification and drug resistance by transient hypoxia. Numerous subclones of Chinese hamster ovary cells selected by growth in Adriamycin or methotrexate-containing medium express elevated levels of GSH per cell. The method was extended to quantitate the GSH content of cells excised from EMT-6/SF mouse tumors that had been treated in vivo with L-buthionine-S-R-sulfoximine, an inhibitor of GSH synthesis. The bivariate analysis (forward angle light scatter versus monochlorobimane fluorescence) of cells derived from these tumors gave excellent resolution of normal and tumor cells and demonstrated extensive heterogeneity in the tumor cell population with respect to GSH content per cell.

Radiation pneumonitis following combined modality therapy for lung cancer: analysis of prognostic factors.

PURPOSE: To identify factors associated with radiation pneumonitis (RP) resulting from combined modality therapy (CMT) for lung cancer. MATERIALS AND METHODS: Series published before 1994 that used CMT for the treatment of lung cancer and explicitly reported the incidence of RP are the basis for this analysis. Factors evaluated included the radiation dose per fraction (Fx), total radiation dose, fractionation scheme (split v continuous), type of chemotherapy and intended dose-intensity, overall treatment time, histology (small-cell lung cancer [SCLC] v non-small-cell lung cancer [NSCLC]), and treatment schedule (concurrent v induction, sequential, or alternating CMT). RESULTS: Twenty-four series, including 27 treatment groups and 1,911 assessable patients, met our criteria for inclusion in this analysis. The median total dose of radiation used in the trials analyzed was 50 Gy (range, 25 to 63 Gy). The median daily Fx used was 2.0 Gy (range, 1.5 to 4.0 Gy). Nineteen series included 22 treatment groups and 1,745 patients treated with single daily fractions. Among these patients, 136 received a daily Fx greater than 2.67 Gy. Five series used twice-daily radiotherapy and included 166 patients (Fx, 1.5 to 1.7 Gy). The incidence of RP was 7.8%. In a multivariate analysis, only daily Fx, number of daily fractions, and total dose were associated with the risk of RP (P < .0001, P < .018, and P < .003, respectively). CONCLUSION: In this analysis, the use of Fx greater than 2.67 Gy was the most significant factor associated with an increased risk of RP. High total dose also appears to be associated with an increased risk, but twice-daily irradiation seems to reduce the risk expected if the same total daily dose is given as a single fraction. High-Fx radiotherapy should be avoided in patients who receive CMT with curative intent.

Stereotactic radiosurgery and radiotherapy: new developments and new directions.

Stereotactic irradiation is a precise method for the delivery of focused radiation beams to small intracranial targets. Treatment can be administered in single or multiple fractions (radiosurgery or stereotactic radiotherapy, respectively). The technology has evolved rapidly because of advances in both hardware and software design. Clinical indications are unfolding through the prospective trial mechanism.

Protection against cis-dichlorodiammine Pt (II) cytotoxicity in vitro by cysteamine.

EMT6/SF cells exposed to cis-dichlorodiammine Pt(II) (DDP) for 1 hr in the presence of 10 mM cysteamine (MEA) survived better than cells treated with DDP alone (DMF-3.1). MEA added to cultures after removal of DDP also protected, even when the two exposures were separated by as long as 4 hr. Protection by MEA was concentration dependent: no significant protection was observed below 1 mM and maximum protection required 5 mM. When cells were incubated with 10 mM MEA immediately after removal of DDP, survival increased as a function of the length of exposure to MEA, reaching a maximum at 2 hr.

Effects of glutathione depletion by buthionine sulfoximine on the sensitivity of EMT6/SF cells to chemotherapy agents or X radiation.

The effects of depletion of cellular glutathione (GSH) on the sensitivity of cultured EMT6/SF cells to chemotherapy agents or x rays under hypoxic and aerated conditions were investigated. Buthionine sulfoximine (BSO), a potent inhibitor of the enzyme gamma-glutamyl-cysteine synthetase, was used to deplete cellular GSH. Addition of BSO (50 microM) to EMT6/SF cultures depleted cellular GSH with a half-time of approximately 2 hr. Cellular GSH reached very low levels within hours of addition of BSO. After removal of BSO, cellular GSH recovered with approximately the same kinetics as was seen for depletion. Incubation of EMT6/SF cells with BSO concentrations of up to 1 mM did not reduce the viability or inhibit growth when exposure was limited to times less than 24 hr. However, for longer exposure times, toxicity and growth inhibition were demonstrated in a dose dependent fashion. EMT6/SF cells were treated with chemotherapy agents under either aerated or extremely hypoxic conditions. Cells were more sensitive to cis-dichlorodiammino Pt(II) (DDP), mitomycin C (MitC), L-phenylalanine mustard (L-PAM), and nitrogen mustard (HN2) when treatment was under hypoxic conditions. The magnitude of this sensitization under hypoxic conditions ranged from a dose modifying factor (DMF) of 1.4 (HN2) to 4.1 (MitC), measured at the 0.1 level of cell survival. Hypoxic EMT6/SF cells were more resistant to the cytotoxic effects of actinomycin D (ActD) under hypoxic conditions (DMF = 10 at SF = 0.3). When cellular GSH was depleted to less than 5% of control by treatment with 50 microM BSO for 12-14 hr, cells were sensitized to DDP, L-PAM and HN2 under both aerated and hypoxic conditions. DMF's ranged from 1.4-6.5, depending on the agent. Hypoxic cell sensitization was never significantly greater than that seen in aerated cells, as was the case for X radiation (DMF = 1.3 for hypoxic cells only). GSH depletion also sensitized to MitC, but only under aerated conditions (DMF = 2.1). Hypoxic EMT6/SF cells were not sensitized to MitC by depletion of GSH. GSH depletion afforded slight protection against ActD toxicity under both aerated and hypoxic conditions. These studies suggest that cellular GSH plays an important role in modifying cellular response to cytotoxic drugs. GSH depletion may sensitize tumor cells to some chemotherapy agents, but differential sensitization of tumors compared to normal tissues, based on hypoxic tumor cells as targets, would not be expected based on these in vitro experiments.

Enhancement of EMT6/SF tumor cell killing by mitomycin C and cyclophosphamide following in vivo administration of buthionine sulfoximine.

We have examined the effect of L-buthionine sulfoximine (BSO) treatment on the subsequent cytotoxicity of mitomycin C (Mit C) and cyclophosphamide (CYC) in EMT6/SF tumors growing in Balb/c mice. GSH content in tumors decreased to a minimum level (less than 30% of control) 12 hr after a single injection of 5 mmole/kg of BSO. The recovery of tumor GSH was protracted and did not reach control levels up to 24 hr following BSO administration. Mit C and CYC were administered 12 hr after BSO injection. Tumor cell survival reached almost the same minimum level (SF = 0.050-0.06%) 1 hour after Mit C (10 mg/kg) treatment with or without BSO. However, 24 hr after Mit C treatment, survival had increased 36-fold (SF = 1.9%) in non-BSO-treated tumors compared to only 4-fold (SF = 0.3%) in the BSO-treated tumors. A dose modifying factor (DMF) of 1.4-1.8 was observed for BSO-pretreated tumors assayed 24 hr after Mit C administration. In CYC-treated tumors, minimum surviving fractions were found 4 hr after CYC treatment with or without BSO treatment. These minimum survival levels were almost the same in BSO-treated tumors following 100 mg/kg CYC (SF = 0.03%) as in control tumors treated with 150 mg/kg CYC (SF = 0.04%). The same increase in cell survival was observed when tumor excision and assay were delayed 24 hr following CYC administration with or without prior BSO treatment. BSO treatment enhanced the cytotoxicity of CYC by DMFs of 1.6-1.9 based on the 24 hr excision assay. Depletion of tumor GSH by BSO caused enhancement of tumor cell killing by the two chemotherapy agents studied, either by increasing cytotoxicity, in the case of CYC or by decreasing PLD recovery following Mit C.

Repair of radiation induced damage--dependence on oxygen and energy status.

Repair of radiation-induced sublethal damage by Chinese hamster V-79 cells is studied under conditions of different pO2 (15 ppm to 21%) and cellular energy status. Cellular ATP content and energy charge are drastically reduced when cells are deprived of both oxygen and nutrient. Both indices are within normal range when cells are provided with nutrients or trace levels of oxygen (300 ppm). When deprived of nutrient, hypoxic (15 ppm O2) cells do not repair SLD, while cells in 300 ppm O2 do. Thus, cellular SLD repair appears to be dependent on cellular energy status which in turn is sensitive to oxygen concentration. Relatedly, nutrient deprived hypoxic cells are sensitized to radiation with storage under 23 degrees C, a phenomenon which may stem from a decrease of endogenous glutathione content.

Radiation therapy with concomitant continuous infusion cisplatin for unresectable nonsmall cell lung carcinoma.

PURPOSE: This pilot study was designed to test the tolerance and effectiveness of concurrent continuous infusion cisplatin and radiotherapy in the treatment of unresectable nonsmall cell lung (NSCL) cancer. METHODS AND MATERIALS: Between July 1989 and July 1991, 92 consecutive patients with either medically or technically inoperable NSCL cancer were treated with thoracic radiotherapy and concomitant chemotherapy. Radiotherapy consisted of a total dose of 70 Gy delivered in 2 Gy daily fractions over 9 weeks with a planned 2-week break after 40 Gy. During the second week of each cycle of radiotherapy, cisplatin was administered, 20 mg/m2/day for 5 days as a continuous infusion. Eighty-five patients were evaluable. RESULTS: Overall response rate was 81.7% (65.9% complete response). Medically operable patients were considered for curative surgical resection following 40 Gy and one cycle of chemotherapy; 11 patients underwent resection with 3/11 having no pathologic evidence of tumor. Median survival for all 85 patients was 11.4 months with a median follow-up of 27 months. Overall survival was 48.2%, 27.5%, and 25% at 12, 24, and 36 months, respectively. Survival was independent of tumor stage, histology and grade, and patient age and gender. Patients having a complete response (n = 54) had a 2-year survival of 42.1% compared to 3.2% for partial-responders and nonresponders (n = 31; p < 0.0001). Patients undergoing surgical resection (n = 11) had a 2-year survival of 75.8% compared to 20.6% for those treated with chemoradiotherapy alone (n = 74). Forty-eight patients have died of their disease. There were two treatment-related deaths, seven deaths of intercurrent disease and three of unknown causes. Eighteen of 25 patients alive at the time of analysis were without evidence of disease. Actuarial local control was 50.6% at 1 year, and 33.3% at 2 years. The distant failure rate was 47.8% at 2 years. Major acute toxicities, mainly hematologic or gastrointestinal, occurred in less than 10% of patients. Esophagitis was mild and infrequent (8.4%). Severe late pulmonary fibrosis occurred in 5.2% of patients and resulted in two treatment-related deaths. CONCLUSION: Concomitant chemoradiotherapy was well tolerated, resulted in a high rate of local control, and in a survival benefit for patients demonstrating a complete response or going on to surgical resection. The incidence of distant metastases continues to be high and future strategies should be directed at improving systemic therapy.

Hyperfractionated radiation therapy for gliomas of the brainstem in children and in adults.

Between February 1984 and September 1990, 60 patients with brainstem gliomas were treated with hyperfractionated radiotherapy in the Department of Radiation Oncology at the University of California, San Francisco. Forty-one children (< or = 18 years) and 19 adults were treated with 100 cGy twice daily with 4-8 hr between doses. Thirty-one patients (21 children and 10 adults) received total doses of 66-72 Gy and 29 patients (20 children and nine adults) received 74-78 Gy. Median follow-up was 208 weeks for all patients (214 weeks for children, 157 weeks for adults). Twenty-three patients (14 children and nine adults) were alive at the time of analysis, surviving 59-359 weeks following treatment. Median actuarial survival was 73.6 weeks overall (72 weeks for children, 190 weeks for adults; p = 0.43). Survival at 12 and 24 months was 65% and 38%, respectively (63% and 32%, for children; 68% and 53% for adults). All patients had pretreatment magnetic resonance imaging by which tumors were classified as either focal or diffuse. No significant pretreatment prognostic factors for adults were identified. In children, significant favorable prognostic factors on univariate analysis were older age (p = 0.001), tumor location in thalamus or midbrain (p = 0.002), focal appearance on MRI scan (p < 0.001) and duration of symptoms > 2 months prior to treatment (p < 0.001). Thirty-five patients had tumor biopsies, leading to a diagnosis in 33 (22 children and 11 adults). Children with moderately anaplastic astrocytomas survived significantly longer than those with glioblastoma multiforme or unbiopsied tumors (p < 0.001). Only duration of symptoms > 2 months remained significant as a favorable prognostic indicator for children on multivariate analysis (p < 0.001). Survival was not significantly different for patients receiving < or = 72 Gy and those receiving > 72 Gy (p = 0.18). No subgroup of patients showed significantly better survival with the higher dose. These findings indicate that hyperfractionated radiotherapy is effective treatment for adults and a subgroup of better prognosis children with brainstem gliomas. There is a subgroup of pediatric patients with extremely poor prognosis for whom even this aggressive treatment does little to extend survival. We conclude that there is no benefit to increasing total dose above 72 Gy for any of the groups analyzed.

Fractionated stereotactic radiotherapy for the treatment of acoustic neuromas: preliminary results.

PURPOSE: To evaluate the efficacy and toxicity of fractionated, stereotactic radiotherapy (SRT) for acoustic neuromas. METHODS AND MATERIALS: Twelve patients with acoustic neuroma were treated with SRT between June 1992 and October 1994. Follow-up ranged from 16-44 months. Patient age ranged from 27-70 (median: 45). Eight patients were treated with primary SRT and four patients were treated after primary surgical intervention for recurrent [3] or persistent [1] disease. Tumor volumes were 1.2-18.4 cm3 (median: 10.1 cm3). Collimator sizes ranged from 30-50 mm (median: 37.5). Tumors received 1.8 Gy/day normalized to the 95% isodose line. Patients received a minimum prescribed dose of 54 Gy in 27-30 fractions over a 6-week period. RESULTS: After a median follow-up of 26.5 months, local control was obtained in 12 out of 12 lesions. Tumor regression was noted in three patients, and tumor stabilization was found in the remaining nine patients. No patient developed a new cranial nerve deficit. One patients developed worsening of preexisting Vth cranial neuropathy and another experienced a decrease in hearing. However, all nine patients with useful hearing prior to SRT maintained useful hearing at last follow-up. CONCLUSIONS: Stereotactic radiotherapy provided excellent local control without new cranial nerve deficits. These results must be viewed as tentative in nature because of the small number of patients and the short median follow-up period.

Intracranial germinoma: the case for lower dose radiation therapy.

PURPOSE: To determine the optimal dose and treatment outcome of patients treated with radiation for intracranial germinoma. METHODS AND MATERIALS: Between 1975 and 1995, 40 patients with the diagnosis of intracranial germinoma were treated with radiation (RT) to the central nervous system. All patients received whole-brain (WB) RT (median dose: 32.4 Gy, range: 15-44.37 Gy) and a boost to the tumor volume (median total tumor volume dose: 52 Gy, range: 45-59.5 Gy). Thirty patients received RT to the spine (median dose: 26 Gy, range: 18.75-37.5 Gy). Four patients were treated with cisplatin-based chemotherapy and WB RT with a boost to the tumor volume (dose range: 51-54 Gy). A low-dose RT only group was defined as < or = 25.5 Gy to the WB (9 patients); < 50 Gy to the primary site (14 patients); and < 22 Gy to the spine (9 patients). Seventeen tumors were biopsy-proven germinoma, and 17 patients presented with multiple midline germinomas (MMG). Among 26 patients who had tumor markers measured, 27% had elevation of beta-human chorionic gonadotropin and by definition, no patient had an elevation of AFP. Twenty-four percent of 26 patients who had spine imaging or cerebral spinal fluid cytology had evidence of tumor seeding at diagnosis. The male to female ratio was 1.9:1. Median age at diagnosis was 14 years for male patients and 9.5 years for female patients (p = 0.02), (overall age ranges: 0.5-31 years). Median follow-up was 62 months (range: 3-226 months). Late effects of 29 patients with follow-up of > or = 20 months and adequate documentation in their medical records were analyzed. RESULTS: The 5-year actuarial rate of disease-free survival (DFS) and overall survival (OS) for biopsy-proven germinomas and presumed germinomas was 97%. No patient died of germinoma. There were no local failures regardless of the dose of RT, elevation of HCG tumor marker, or CSF dissemination at presentation. At presentation 22 patients had evidence of at least one endocrine abnormality. At follow-up there were no new patients diagnosed with an endocrine abnormality; however, 13 out of 22 patients had an increase in the number of endocrine deficiencies requiring hormone replacement. At presentation, 14 patients showed evidence of growth retardation. At follow-up there were no new cases of growth failure in the remaining patients. CONCLUSIONS: Germinomas are highly curable with RT alone. Lower doses of RT to the craniospinal axis without chemotherapy appear to produce equally effective DFS and OS as do higher doses of RT or combination chemotherapy and RT. Craniospinal RT may be indicated for patients with MMG or patients with evidence of spinal seeding. Long-term effects of growth retardation, and other endocrine deficiencies appear to be correlated with disease at presentation rather than solely with treatment.

Beam shaping for conformal fractionated stereotactic radiotherapy: a modeling study.

PURPOSE: The patient population treated with fractionated stereotactic radiotherapy (SRT) is significantly different than that treated with stereotactic radiosurgery (SRS). Generally, lesions treated with SRT are larger, less spherical, and located within critical regions of the central nervous system; hence, they offer new challenges to the treatment planner. Here a simple, cost effective, beam shaping system has been evaluated relative to both circular collimators and an ideal dynamically conforming system for effectiveness in providing conformal therapy for these lesions. METHODS AND MATERIALS: We have modeled a simple system for conformal arc therapy using four independent jaws. The jaw positions and collimator angle are changed between arcs but held fixed for the duration of each arc. Eleven previously treated SRT cases have been replanned using this system. The rectangular jaw plans were then compared to the original treatment plans which used circular collimators. The plans were evaluated with respect to tissue sparing at 100%, 80%, 50%, and 20% of the prescription dose. A plan was also done for each tumor in which the beam aperture was continuously conformed to the beams eye view projection of the tumor. This was used as an ideal standard for conformal therapy in the absence of fluence modulation. RESULTS: For tumors with a maximum extent of over 3.5 cm the rectangular jaw plans reduced the mean volume of healthy tissue involved at the prescription dose by 57% relative to the circular collimator plans. The ideal conformal plans offered no significant further improvement at the prescription dose. The relative advantage of the rectangular jaw plans decreased at lower isodoses so that at 20% of the prescription dose tissue involvement for the rectangular jaw plans was equivalent to that for the circular collimator plans. At these isodoses the ideal conformal plans gave substantially better tissue sparing. CONCLUSION: A simple and economical field shaping device has been shown to provide all of the beam shaping advantage of a hypothetical ideal dynamically conforming system at the prescription level. This system may be immediately implemented in the clinic. It offers a substantial advantage over the currently used circular collimators in the high dose region with equivalent performance in the low dose region.

Childhood optic chiasm gliomas: radiographic response following radiotherapy and long-term clinical outcome.

PURPOSE: In children with chiasmal gliomas, radiation therapy can arrest progressive visual and neurologic impairment. We examined the radiographic response and clinical outcomes after irradiation. METHODS AND MATERIALS: Forty-two children (median age at diagnosis, 6.6 years) with chiasmal gliomas were managed as follows: 11 asymptomatic patients with neurofibromatosis-1 (NF-1) were observed only; 2 patients, less than 3 years old, underwent surgery and chemotherapy to delay irradiation; and 29 patients with progressive disease received radiation with or without prior surgery or chemotherapy. Time to radiographic response, long-term tumor control and late sequelae were reviewed for the 29 irradiated patients. RESULTS: The probability of at least 50% radiographic response at 24 months after irradiation was 18.1% and increased to 38.2% by 48 months and 45.9% by 60 months. By actuarial analysis, the median time for such radiographic response was 62 months. For the 29 irradiated patients, the 10-year freedom from progression and overall survival rates were 100% and 89%, respectively (median follow-up for surviving patients, 108 months). Stabilization or improvement in vision occurred in 81% of 26 evaluable irradiated patients. CONCLUSIONS: Notable radiographic response may be observed years after irradiation. Radiation therapy provides excellent long-term tumor control and vision preservation or improvement in the majority of patients with progressive chiasmal gliomas.

Glutathione depletion in tissues after administration of buthionine sulphoximine.

Buthionine sulphoximine (BSO) an inhibitor of glutathione (GSH) biosynthesis, was administered to mice in single and repeated doses of 0.5, 1 and 5 mmol kg-1 (i.p.). The resultant pattern of GSH depletion was studied in liver, kidney, skeletal muscle and three types of murine tumor. Liver and kidney exhibited a rapid depletion to GSH levels of ca. 20% of controls after single doses of 1-5 mmol kg-1 BSO. Muscle was depleted to a similar level, but at a slower rate after a single dose. All three tumors required repeated administration of BSO over several days to obtain a similar degree of depletion to that shown in the other tissues.