In vitro hypoxic cytotoxicity and hypoxic radiosensitization. Efficacy of the novel 2-nitroimidazole N,N,N-tris[2-(2-nitro-1H-imidazol-1-yl)ethyl]amine.

BACKGROUND AND PURPOSE: Tumor hypoxia is a major problem in radiation therapy of solid tumors because of the radiosensitizing effect of oxygen. Nitroimidazole-containing compounds are oxygen mimetics accumulating in hypoxic tumor areas. However, the broad use of 2-nitroimidazoles as a hypoxic radiosensitizer is limited by their partially low efficacy and/or high neurotoxicity. MATERIALS AND METHODS: Here, we characterized the in vitro hypoxic cytotoxicity and hypoxic radiosensitizing efficacy of N,N,N-tris [2-(2-nitro-1H-imidazol-1-yl)ethyl]amine (PRC) in a hypoxia-sensitive lymphoma and a hypoxia-resistant glioblastoma cell line by colony formation assay and flow cytometry. RESULTS: PRC exerted high hypoxic cytotoxic and radiosensitizing action on both cell lines at almost absent toxicity under normoxic conditions. In particular, under hypoxia, but not normoxia, PRC targeted the mitochondria resulting in oxidative stress, G(2)/M cell cycle arrest, and triggering of the intrinsic apoptosis pathway. CONCLUSION: Our in vitro findings suggest that PRC might be a promising new 2-nitroimidazole for improving radiation therapy of hypoxic tumors in vivo.

Are there biologic differences between male and female breast cancer explaining inferior outcome of men despite equal stage and treatment?!

BACKGROUND: Reasons for inferior outcome of male compared to female breast cancer are still under debate. Therefore, we retrospectively analyzed male breast cancer cases to figure out possible treatment- and gender-related differences. PATIENTS AND METHODS: A total of 40 men (median age 62 years) were curatively treated with mastectomy and postoperative radiotherapy from 1982-2007. They presented predominantly in stages II and IIIb. Postoperative radiotherapy was applied with doses of 1.8-2.5 Gy to a median of 50 Gy including regional lymphatics in 22 patients. Adjuvant systemic treatment consisted of chemotherapy (22.5%) and antihormonal treatment (55%). For reasons of comparison, we estimated outcome of a virtual female matched cohort for no/equal to men/optimal adjuvant treatment with the Adjuvant!Online(®) 8.0 algorithm. RESULTS: After a median follow-up of 47 months, the estimated 5-year local control rate was 97%, disease-free and distant metastasis-free survival rates reached 79% and 82%, respectively. With update of survival data by tumor registry, mean overall survival reached 120 months with 5- and 10-year overall survival rates of 66% and 43%, respectively. Predominant prognostic factor was T-stage for overall survival (T1/2 vs. T4: > 80% vs. 30%). The generated virtual matched cohorts of women with equal characteristics reached superior 10-year-overall survival for no/equal to men/optimal adjuvant treatment with 55/59/68%. CONCLUSION: Compared to historical and virtual matched cohorts of women, male breast cancer patients had inferior outcome despite of equal stage and treatment which indicates that biological differences (of tumor or population) may contribute to worse prognosis.

Adjuvant radiochemotherapy in patients with locally advanced high-risk cervical cancer.

BACKGROUND AND PURPOSE: The aim of this retrospective study was to analyze the outcome of patients with locally advanced cervical carcinoma treated by adjuvant radiochemotherapy and to determine risk factors for local and distant relapse. Furthermore, acute and late effects of treatment were recorded. PATIENTS AND METHODS: A total of 72 patients with FIGO stages I-III cervical carcinoma were treated by radical hysterectomy, pelvic lymphadenectomy, and postoperative radiochemotherapy. Only patients with positive pelvic lymph nodes, parametrial involvement, positive margins, or tumor bulk were eligible. Patients were irradiated with a standard pelvic field (50.4 Gy in 28 fractions). The majority of patients received platinum-based chemotherapy. RESULTS: After a median follow-up of 37 months, estimated 1-, 2-, and 4-year disease-free survival (DFS) and overall survival (OS) rates were 89%, 80%, 68% and 95%, 88%, 76%, respectively. Nine of the 72 patients had pelvic recurrences including only 1 isolated local failure; 23 of the 72 patients presented with distant relapse. The majority of relapses occurred within the first 3 years after adjuvant treatment. The number of positive pelvic lymph nodes (> 1) was the strongest prognostic factor for DFS. Treatment was well tolerated with transient acute hematologic (~30%) and gastrointestinal (~30%) grade 3 toxicity. Small bowel obstruction (~6%) was the only important late sequelae. CONCLUSION: Adjuvant radiochemotherapy in patients with advanced cervical cancer and several risk factors is highly effective to prevent local relapse. Future efforts to improve outcome should be placed on improvement of systemic control especially in subgroups with high-risk features for distant relapse. Combined treatment was well tolerated with moderate acute and late toxicity.

Postoperative versus definitive chemoradiation in early-stage anal cancer. Results of a matched-pair analysis.

BACKGROUND AND PURPOSE: The goal of the present study was to comparatively assess the results of definitive chemoradiation (CRT) with or without previous macroscopically complete resection in patients with early-stage node-negative (T1-2 N0) anal carcinoma. PATIENTS AND METHODS: A total of 20 patients with T1-2 N0 anal carcinoma who received radiotherapy (RT) with or without chemotherapy following incidental R0/1 tumor resection (S/CRT group) were selected. These were matched to 20 comparable patients who underwent definitive chemoradiation without previous surgery (CRT group). Major objectives of this analysis were treatment outcomes in terms of locoregional tumor control (LRC), overall survival (OS), colostomy-free survival, and toxicity. RESULTS: Patients treated postoperatively received significantly lower RT doses (median 54.0 Gy vs. 59.7 Gy; p < 0.001) and less frequently concomitant chemotherapy than those treated definitely. The 5-year LRC and 5-year OS rates were 97.5% and 90.0%, respectively, without significant differences between the S/CRT and the CRT groups. The distribution of acute and late toxicities was comparable, and the 5-year colostomy-free survival was 95% in both groups. CONCLUSION: This matched-pair comparison of incidental R0/1 resection plus dose-reduced CRT with standard definitive CRT of early-stage anal cancer shows similar treatment results. Thus, dose-reduced RT with or without chemotherapy may be considered in R0/1 resected patients with T1-2 N0 anal carcinoma.

Limited disease of extra-pulmonary small cell carcinoma. Impact of local treatment and nodal status, role of cranial irradiation.

PURPOSE: As extra-pulmonary small cell carcinoma (EPSCC) is a rare entity of tumors, the available treatment recommendations are mainly based on retrospective analyses and deduction from treatment of small cell lung cancer. The aim of this study was to provide a detailed analysis concerning prognostic factors and treatment modalities. PATIENTS AND METHODS: A total of 20 patients with limited disease (LD) of EPSCC treated at our institution from 1999­2009 were retrospectively analyzed. Data were gathered from chart review. Localization, lymph node involvement, as well as local and systemic treatment were documented and their impact on pattern of failure and survival times statistically evaluated. RESULTS: With a median follow-up of 21 months, the estimated median overall- and disease-free survival were 59 and 25 months, respectively. Local control was excellent with 100% at 2 years. Nodal involvement was observed in 74% (n = 14/19) of evaluable patients. However, outcome was not altered by this parameter. Local treatment consisted of surgery in 10 cases, radiotherapy in 7 cases, and a combination of both in 3 cases. Only 3 patients (15%) developed hematogenous central nervous system metastases, while none of the patients received prophylactic cranial irradiation. CONCLUSION: Nodal involvement did not worsen prognosis. Local control was excellent irrespective of local treatment modality and the leading cause of failure was distant metastasis. Therefore, systemic treatment should not be omitted. Prophylactic cranial irradiation might be dispensable but discussed for head and neck malignancies.

Gene expression analysis in chronic postradiation proctopathy.

PURPOSE: Radiotherapy is one of the important treatment modalities for tumors of pelvic organs. The fixed location of the rectum and its anatomic relationship with other pelvic organs makes it prone to radiation injury resulting in chronic radiation proctopathy in 5% to 20% of patients. Endothelial dysfunction has been associated with a number of pathophysiological processes. Endothelial cells synthesize and release various factors that regulate angiogenesis, inflammatory responses, hemostasis, as well as vascular tone and permeability. METHODS: Rectum tissue samples from 20 patients with established chronic radiation proctopathy were analysed for the expression of genes related to oxidative stress, tissue hypoxia, angiogenesis, and inflammation [endoglin (ENG), activin receptor-like kinase 1 (ALK1), platelet endothelial cell adhesion molecule 1 (PECAM), vascular endothelial growth factor (VEGF), fibroblast growth factor 2 (FGF2), hypoxia-inducible factor 1 (HIF-1), and interleukin-1 beta (IL-1ß)]. RESULTS: Overexpression of HIF-1, VEGF, FGF2, and IL-1ß was detected in affected tissue. For the first time, a significant suppression of activin receptor-like kinase 1 and ENG could be revealed. CONCLUSION: The data provided here allow further insight into the pathogenesis of radiation-induced rectum injury. Radiation-induced damage is not confined to a single event but involves complex signaling between different pathways, enhancing and maintaining the processes that lead to mucosal damage. The results indicate that postradiation tissue hypoxia is critical for fibrosis, which involves changes in the expression of profibrotic and angiogenic factors in rectal tissue.

Tumour hypoxia: impact on biology, prognosis and treatment of solid malignant tumours.

Tumour hypoxia is a major constraint for radiotherapy and many types of chemotherapy. A variety of different pathogenetic mechanisms contribute to the development of hypoxia in solid tumours. Hypoxia is associated with unfavourable prognosis, regardless of the treatment modality applied. Two different effects have been considered to explain the deleterious effects of hypoxia on the outcome of tumour patients. The first aspect encompasses the direct interference of hypoxia with antineoplastic treatment modalities. The efficacy of ionizing radiation, but also of a variety of cytotoxic drugs and cytokines rely directly on adequate oxygen tensions. The second aspect concerns the effects of hypoxia on the biology of tumour and stromal cells. Hypoxia is related to malignant progression, increased invasion, angiogenesis and an increased risk of metastasis formation. Possibly, hypoxia is furthermore a stressor which selects cells with increased resistance to apoptosis and thereby indirectly contributes to treatment resistance. This article reviews in brief the specific pathophysiology of tumour oxygenation and its implications for prognosis, tumour treatment and biology.

Expression pattern of the urokinase-plasminogen activator system in rat DS-sarcoma: role of oxygenation status and tumour size.

The urokinase plasminogen activator system plays a central role in malignant tumour progression. Both tumour hypoxia and enhancement of urokinase plasminogen activator, urokinase plasminogen activator-receptor and plasminogen activator inhibitor type 1 have been identified as adverse prognostic factors. Upregulation of urokinase plasminogen activator or plasminogen activator inhibitor type 1 could present means by which hypoxia influences malignant progression. Therefore, the impact of hypoxia on the expression pattern of the urokinase plasminogen activator system in rat DS-sarcoma in vivo and in vitro was examined. In the in vivo setting, tumour cells were implanted subcutaneously into rats, which were housed under either hypoxia, atmospheric air or hyperoxia. For in vitro studies, DS-sarcoma cells were incubated for 24 h under hypoxia. Urokinase plasminogen activator and urokinase plasminogen activator-receptor expression were analysed by flow cytometry. Urokinase plasminogen activator activity was measured using zymography. Plasminogen activator inhibitor type 1 protein levels in vitro and in vivo were examined with ELISA. PAI-1 mRNA levels were determined by RT-PCR. DS-sarcoma cells express urokinase plasminogen activator, urokinase plasminogen activator-receptor, and plasminogen activator inhibitor type 1 in vitro and in vivo. The urokinase plasminogen activator activity is enhanced in DS-sarcomas compared to normal tissues and rises with increasing tumour volume. The oxygenation level has no impact on the urokinase plasminogen activator activity in cultured DS-sarcoma cells or in solid tumours, although in vitro an increase in plasminogen activator inhibitor type 1 protein and mRNA expression after hypoxic challenge is detectable. The latter plasminogen activator inhibitor type 1 changes were not detectable in vivo. Hypoxia has been demonstrated to contribute to the upregulation of some components of the system in vitro, although this effect was not reproducible in vivo. This may indicate that the serum level of plasminogen activator inhibitor type 1 is not a reliable surrogate marker of tumour hypoxia.

Clinical indications and biological mechanisms of splenic irradiation in autoimmune diseases.

BACKGROUND: Splenic irradiation (SI) is a fairly unknown treatment modality in autoimmune disorders like autoimmune thrombocytopenia (AIT) or autoimmune hemolytic anemia (AIHA), which may provide an effective, low toxic and cost-effective treatment for selected patients. PATIENTS, MATERIALS AND METHODS: This article reviews the limited experiences on splenic irradiation in autoimmune thrombocytopenia by analyzing the current studies including 71 patients and some preliminary reports on splenic irradiation in autoimmune hemolytic anemia. RESULTS: In autoimmune thrombocytopenia between 40 and 90% of all patients responded, but most of them relapsed within 4 to 6 months after splenic irradiation. Between 10 and 20% of all patients had a sustained response. The efficacy of splenic irradiation in HIV-associated cases of thrombocytopenia is probably lower than in other forms of autoimmune thrombocytopenia, but especially in this group immunosuppressive drug treatment of autoimmune thrombocytopenia exposes some problems. In autoimmune hemolytic anemia there are some case reports about efficacy of splenic irradiation. Toxicity of splenic irradiation in both diseases was very moderate. CONCLUSIONS: For HIV patients, for elderly patients or patients at high risk for complications following splenectomy splenic irradiation might be a treatment option. Splenic irradiation as preoperative treatment in patients not responding to or not suitable for immunosuppressive drugs prior to splenectomy may be a promising new application of splenic irradiation to reduce adverse effects of splenectomy in thrombocytopenic patients. A further analysis of the biological mechanisms underlying splenic irradiation may help to improve patient selection, to optimize dose concepts and treatment schedules and will improve understanding of radiotherapy as an immunomodulatory treatment modality.

Clinical indications and biological mechanisms of splenic irradiation in chronic leukaemias and myeloproliferative disorders.

Splenic irradiation (SI) was the first efficient treatment for chronic leukaemia, but with the emergence of effective drugs its use has been more and more restricted to advanced cases presenting with splenomegaly. But in selected patients who are not responsive or not suitable to drug treatment, SI may offer still an effective, low toxic and cost-effective palliative modality. Eight studies of SI in chronic lymphatic leukaemia (CLL) including 198 patients, six reports about SI in prolymphocytic leukaemia (PLL), including 18 patients, one study and six case reports about SI in hairy cell leukaemia (HCL) and nine studies about SI in myeloproliferative disorders has been analyzed. In CLL, symptoms of splenomegaly have been improved in 50-87% of all patients with overall doses between 4 and 10 Gy in mostly 1-Gy fractions. PLL seems to be more resistant to SI with a median response rate of 66%. Casuistic reports described also efficacy of SI in HCL patients using similar radiation schedules. Symptomatic relief is also provided by SI in myeloproliferative disorders using lower overall doses between 1 and 9 Gy with small single fractions of 0.25 Gy (median). Acute toxicity was low in lymphoid disorders, but higher in myeloproliferative disorders with severe cytopenia in 10-30% of all cases, indicating the need for a cautious fractionation schedule. Interestingly, even complete systemic remissions after SI in all types of lymphoproliferative disorders have been described. Different mechanisms underlying SI such as direct cell kill, immune modulation via changes in lymphocyte subsets or cytokine induction or "radiotherapeutic" splenectomy with high doses are discussed.

Stress-induced hormonal alterations.

The hormonal alterations observed during critical illness are extremely diverse, being both adaptive and maladaptive. Central and peripheral mechanisms are at play depending on the duration and severity of the insult. Perturbations in the endocrine axes are exacerbated by endogenous mediators and by pharmacologic intervention. The emergence of the concept of the wound, or inflammatory lesion, as an endocrine organ introduces new potential mechanisms of stress physiology and possible interventional strategies. Whether the physiology of the wound can be manipulated to heighten organism resistance to stress either emergently or electively is theoretically appealing and supported by animal models. Currently, however, it remains a subject of ongoing controversy and research.

Biological effects of ionizing radiation on human blood compounds ex vivo.

PURPOSE: Blood compounds are irradiated ex vivo to prevent transfusion-associated graft-versus-host-disease. Recently, ex vivo irradiation of re-transfused wound blood has been proposed to prevent metastatic spread in patients with malignant tumors, an issue requiring different dose concepts. To determine effects on blood cells we examined the impact of various doses of ionizing radiation. METHODS: Full blood was irradiated with doses of 10-150 Gy. Potassium, LDH and hemoglobin levels were determined 2 h-96 h after irradiation. The lymphocyte proliferation after irradiation was measured by means of a lymphocyte-transformation assay. The impact of irradiation on mitogen-induced secretion of cytokines was determined by the ELISA technique, and P-selectin expression as an indicator of platelet activation was analyzed by flow-cytometry. RESULTS: Potassium levels increase with aging and irradiation dose. Mitogenic capacity is reduced by over 90% with moderate doses of 10-20 Gy, but a residual proliferation is still detectable up to 50 Gy. No enhancement of extracellular cytokine levels is detectable, but the cytokine release is reduced by radiation. Neither induction of platelet activation nor abrogation of activation has been detected. CONCLUSIONS: Doses of 30-50 Gy abrogate lymphocyte proliferation almost completely. In this range we did not observe severe adverse effects on blood transfusions. Hemolysis might be enhanced when the samples are stored for a longer period after irradiation.

Soluble levels of CD-95, CD 95-L and various cytokines after exposing human leukocytes to ionizing radiation.

BACKGROUND: Local radiotherapy induces systemic effects and influences the immune system. Some of these effects can hardly be explained by direct cellular reactions, but might be triggered by humoral factors. MATERIALS AND METHODS: In seminoma patients we measured the influence of radiotherapy on various lymphocyte subsets by FACS-analysis. We analysed the release of cytokines, CD95 (Apo-1/Fas) and CD 95-L (Fas-Ligand) of human blood cells after exposure to ionizing radiation in vitro and in vivo by ELISA-technique. RESULTS: Although radiation leads to a differential and sustained reduction of all lymphocytes subsets in vivo no increase of cytokine levels, or of levels of CD95 or CD 95-L could be detected by ELISA in vivo or in vitro. On the contrary mitogen-induced cytokine production in vitro was inhibited by radiation. CONCLUSIONS: Local radiation induced a significant systemic decrease of all lymphocyte subsets, but neither a systemic release of cell death mediators like CD95 or various cytokines, nor a release in the extracellular space in vitro were detectable.

Cation metabolism during propofol sedation with and without EDTA in patients with impaired renal function.

OBJECTIVE: To compare the effects of propofol with and without disodium edetate (EDTA) on cation metabolism in intensive care unit (ICU) patients with renal insufficiency who received propofol or propofol plus EDTA (propofol EDTA) for sedation and mechanical ventilation. DESIGN: Double-blind, randomised, multicentre study. SETTING: Medical and surgical ICUs from 5 hospitals. PATIENTS: Thirty-nine ICU patients with acute and chronic renal impairment expected to require at least 24 hours of continuous sedation and respiratory failure necessitating mechanical ventilation. INTERVENTIONS: Propofol or propofol EDTA administered for sedation by continuous intravenous infusion. MEASUREMENTS AND RESULTS: The depth of sedation, as measured by the Modified Ramsay Sedation Scale, was similar in the 2 groups, when adjusted for dosing differences. The amount of propofol required to maintain adequate sedation was decreased in both groups compared to propofol requirements in ICU patients with normal renal function. EDTA levels were elevated at baseline in both groups. In the propofol EDTA group, the EDTA levels increased further by 20 % but decreased to below baseline EDTA levels at 48 hours after sedation. In the propofol group, EDTA levels decreased during sedation and remained below baseline levels at 48 hours after sedation. PATIENTS in both groups were hypocalcaemic and hyperphosphataemic at baseline with low levels of 1,25-dihydroxyvitamin D and elevated parathyroid hormone (PTH) levels. Other than a slight difference in ionised serum calcium levels at 4 h after the start of sedation, there were no significant differences observed in serum calcium levels between the two groups. There were no significant differences in 1,25-dihydroxyvitamin D or PTH levels over time between the two groups. There was no significant effect on renal function in either group. CONCLUSIONS: The results of this study suggest that adding EDTA to propofol does not adversely affect cation homeostasis or renal function when used for sedation of ICU patients with renal insufficiency. Although EDTA levels increased over time from baseline levels in patients with renal insufficiency who receive propofol EDTA, this increase does not appear to be clinically significant, and EDTA levels return to below baseline levels within 48 hours of discontinuing the propofol EDTA infusion. The efficacy of propofol with and without EDTA also appears comparable in these patients.

Chemotherapy in the treatment of recurrent glioblastoma multiforme: ifosfamide versus temozolomide.

PURPOSE: Despite the progress made in neurosurgery and radiotherapy, the prognosis of glioblastoma multiforme (GB) is poor, due to the lack of an effective salvage therapy. In vitro analysis revealed activity for ifosfamide and temozolomide. The usefulness of these agents in recurrent disease was investigated. METHODS: Six adult patients with recurrent GB received one to four courses of 1,500 mg/m2 ifosfamide given over 5 days intravenously. Furthermore, temozolomide (100-200 mg/m2) was given orally over 5 days to 14 patients. RESULTS: After ifosfamide treatment, one partial response and two cases of stable disease were observed. The median survival time was 24 weeks (range of 9-52 weeks). Toxicity analysis revealed one paranoid reaction, three grade III leukocytopenia, and one grade I-II nausea, anemia, and hematuria. Temozolomide therapy resulted in three partial responses and four cases of stable disease. The median survival time (Kaplan-Meier) was 21 weeks (range 4-64 weeks). The major toxicities were grade I-II nausea and hematological side effects (one case of grade IV leuko- and thrombocytopenia). CONCLUSIONS: Ifosfamide treatment might be a feasible approach, but it necessitates hospitalization. Temozolomide showed promising results. Due to its oral application, the patient's quality of life (time out of hospital) is favorable. Subgroups with improved survival were observed.

Impact of localized radiotherapy on blood immune cells counts and function in humans.

Immune cells subsets were prospectively analyzed after localized radiotherapy (LRT). LRT reduced the levels of all lymphocyte subsets, with B-cells and naive T-cells being most sensitive. Lymphocyte function was suppressed, but still within the normal range. Rapid recovery of cytotoxic T-cells/natural killer cells after LRT and the functional suppression within normal levels explains the low incidence of infections after LRT.

Radiosurgery for the treatment of brain metastases in renal cell carcinoma.

BACKGROUND: In the treatment of brain metastases using a stereotactically modified linear accelerator it could be shown that a single dose between 15 and 25 Gy leads to partial or complete remission of so-called radioresistant metastases from melanoma and hypernephroma. Radiosurgery of brain metastases then started in centers all over the world, however, experiences with brain metastases of renal cell carcinoma are yet limited. The aim of this analysis is therefore to present the treatment results of radiosurgery of brain metastases. Furthermore, in this paper prognostic subgroups shall be defined, in order to establish guidelines for an optimal therapy strategy. MATERIALS AND METHODS: Radiosurgery means stereotactically guided high-precision irradiation methods by extremely focussing ionizing radiation within the target volume as a single dose application. The characteristic steep dose decrease allows the selective destruction of small intracranial lesions, while the surrounding brain tissue is optimally protected. Two methods, Gamma Knife and stereotactic modified linear accelerator are clinically available. RESULTS: In larger studies from different groups all over the world, local tumor control rates from 85% to 95%, recurrence rates from 6% to 15% and side effects between 3% and 15% have been attained, independent of the system used. Prognostic factors, like volume of metastases < 10 ml, applied dose > 18 Gy, one or two metastases, absence of extracranial metastases, good patient performance with a Karnofsky score > 70%, primary treatment and more than one year between primary diagnosis and brain metastases showed a trend toward improved survival. Depending on the prognostic factors the median survival after radiosurgery ranged from 6 to 12 months. Retrospective comparison of radiosurgery and surgical series suggest that both modalities attain similar results. The dose can be applied with an accuracy of 0.3 mm. DISCUSSION: Based on these experiences, brain metastases can be treated by radiosurgery, primarily in patients with one or two metastases or in combination with whole brain irradiation as a boost in patients with more than two metastases. Furthermore with radiosurgery a new treatment modality exists to re-irradiate patients who have been failed after surgery or whole brain irradiation.