A novel epidermal growth factor receptor variant lacking multiple domains directly activates transcription and is overexpressed in tumors.

The epidermal growth factor receptor (EGFR) is essential to multiple physiological and neoplastic processes via signaling by its tyrosine kinase domain and subsequent activation of transcription factors. EGFR overexpression and alteration, including point mutations and structural variants, contribute to oncogenesis in many tumor types. In this study, we identified an in-frame splice variant of the EGFR called mini-LEEK (mLEEK) that is more broadly expressed than the EGFR and is overexpressed in several cancers. Unlike previously characterized EGFR variants, mLEEK lacks the extracytoplasmic, transmembrane and tyrosine kinase domains. mLEEK localizes in the nucleus and functions as a transcription factor to regulate target genes involved in the cellular response to endoplasmic reticulum (ER) stress, including the master regulator of the unfolded protein response (UPR) pathways, molecular chaperone GRP78/Bip. We demonstrated that mLEEK regulates GRP78 transcription through direct interaction with a cis-regulatory element within the gene promoter. Several UPR pathways were interrogated and mLEEK expression was found to attenuate the induction of all pathways upon ER stress. Conversely, knockdown of mLEEK resulted in caspase-mediated cell death and sensitization to ER stress. These findings indicate that mLEEK levels determine cellular responses to unfavorable conditions that cause ER stress. This information, along with the overexpression of mLEEK in tumors, suggests unique strategies for therapeutic intervention. Furthermore, the identification of mLEEK expands the known mechanisms by which the EGFR gene contributes to oncogenesis and represents the first link between two previously disparate areas in cancer cell biology: EGFR signaling and the UPR.

Multimodality treatment with intensity modulated radiation therapy for esophageal cancer.

The objective of this study is to determine the feasibility and report the outcome of patients with locally advanced esophageal cancer treated with preoperative or definitive chemoradiotherapy (CRT) using intensity-modulated radiation therapy (IMRT). Between 2003 and 2007, 30 patients with non-cervical esophageal cancer received concurrent chemotherapy and IMRT at Stanford University. Eighteen patients were planned for definitive CRT and 12 were planned for preoperative CRT. All patients had computed tomography-based treatment planning and received IMRT. The median dose delivered was 50.4 Gy. Patients planned for preoperative CRT underwent surgery 4-13 weeks (median 8.3 weeks) following completion of CRT. Median follow-up of surviving patients from start of RT was 24.2 months (range 8.2-38.3 months). The majority of tumors were adenocarcinomas (67%) and poorly differentiated (57%). Tumor location was 7% upper, 20% mid, 47% lower, and 27% gastroesophageal junction. Actuarial 2-year local-regional control (LRC) was 64%. High tumor grade was an adverse prognostic factor for LRC and overall survival (OS) (P= 0.015 and 0.012, respectively). The 2-year LRC was 83% vs. 51% for patients treated preoperatively vs. definitively (P= 0.32). The 2-year disease-free and OS were 38% and 56%, respectively. Twelve patients (40%) required feeding tube placement, and the average weight loss from baseline was 4.8%. Twelve (40%) patients experienced grade 3+ acute complications and one patient died of complications following feeding tube placement. Three patients (10%) required a treatment break. Eight patients (27%) experienced grade 3 late complications. No grade 4 complications were seen. IMRT was effective and well tolerated. Disease recurrence remains a challenge and further investigation with dose escalation to improve LRC and OS is warranted.

Pathological response after chemoradiation for T3 rectal cancer.

OBJECTIVE: The aim of this study was to investigate the effect of preoperative chemoradiotherapy (CRT) on nodal disease in locally advanced rectal adenocarcinoma. METHOD: Thirty-two patients staged uT3N0 and 27 patients staged uT3N1 rectal adenocarcinoma who underwent pre-CRT staging using endoscopic ultrasound or rectal protocol CT were included. The median radiation dose was 50.4 Gy (range: 45-50.4 Gy) at 1.8 Gy per fraction and all patients received concurrent 5-FU or capecitabine-based chemotherapy. Low anterior resection or abdomino-perineal resection occurred at a median of 46 days (range: 27-112 days) after CRT. RESULTS: Eleven of 32 uT3N0 patients (34.4%) and 13 of 26 uT3N1 patients (50.0%) had ypN+ (P = 0.29). For patients with uT3N0, 10 of 20 (50.0%) with ypT2-3 and 1 of 12 (8.3%) with ypT0-1 were ypN+ (P = 0.02). For patients with uT3N1, 12 of 20 (60.0%) with ypT2-3 and 1 of 6 (16.7%) with ypT0-1 were ypN+ (P = 0.16). Overall, the ypN+ rate was 11.1% in the ypT0-yT1 group compared with 55.0% in the ypT2-yT3 group (P = 003). Among patients with uT3N0 disease, the ypN+ rate in patients who had surgery > 46 days vs 46 days vs 46 days vs

Preventing oxidative stress: a new role for XBP1.

Antioxidant molecules reduce oxidative stress and protect cells from reactive oxygen species (ROS)-mediated cellular damage and probably the development of cancer. We have investigated the contribution of X-box-binding protein (XBP1), a major endoplasmic reticulum stress-linked transcriptional factor, to cellular resistance to oxidative stress. After exposure to hydrogen peroxide (H(2)O(2)) or a strong ROS inducer parthenolide, loss of mitochondrial membrane potential (MMP) and subsequent cell death occurred more extensively in XBP1-deficient cells than wild-type mouse embryonic fibroblast cells, whereas two other anticancer agents induced death similarly in both cells. In XBP1-deficient cells, H(2)O(2) exposure induced more extensive ROS generation and prolonged p38 phosphorylation, and expression of several antioxidant molecules including catalase was lower. Knockdown of XBP1 decreased catalase expression, enhanced ROS generation and MMP loss after H(2)O(2) exposure, but extrinsic catalase supply rescued them. Overexpression of XBP1 recovered catalase expression in XBP1-deficient cells and diminished ROS generation after H(2)O(2) exposure. Mutation analysis of the catalase promoter region suggests a pivotal role of CCAAT boxes, NF-Y-binding sites, for the XBP1-mediated enhancing effect. Taken together, these results indicate a protective role of XBP1 against oxidative stress, and its positive regulation of catalase expression may at least in part account for this function.

Tissue effects after stereotactic body radiotherapy using cyberknife for patients with abdominal malignancies.

AIMS: To report the tissue effects of treatment with single fraction stereotactic body radiotherapy (SBRT) using Cyberknife on malignant tumours of the abdomen and adjacent normal organs. MATERIALS AND METHODS: The data from four autopsies with unresectable pancreatic carcinoma and one lymph node excision from a case of recurrent neuroblastoma were reviewed for radiation-related tissue effects within the primary cancer and the normal organs within the radiation field. RESULTS: Cases of unresectable pancreatic carcinoma consistently showed radiation-induced changes in both the primary tumour and the adjacent, non-malignant colorectal tissue. An additional case of lymph nodes exposed to stereotactic radiation showed typical radiation-related changes, including lymphocyte depletion and capsular fibrosis. CONCLUSIONS: A myriad of radiation-related tissue effects are seen after SBRT with Cyberknife. The changes parallel those reported after conventionally fractionated radiotherapy and suggest that the pathophysiological mechanisms of radiation-induced normal tissue damage are similar for biologically equivalent single and fractionated doses of radiotherapy.

Preoperative versus postoperative radiotherapy for locally advanced gastroesophageal junction and proximal gastric cancers: a comparison of normal tissue radiation doses.

Locoregional relapse occurs in over half of gastric cancer patients who undergo potentially curative resection. Adjuvant chemoradiation reduces locoregional relapse, but often requires irradiating large fields and is limited by poor patient tolerance. This study explores the potential dosimetric benefit in reducing the radiation dose to normal structures by treating gastroesophageal (GE) junction/proximal gastric cancers with preoperative rather than adjuvant radiotherapy. Five cases of GE junction/proximal gastric cancer patients treated postoperatively with curative intent were selected. The actual target contours were then modified to reflect hypothetical target volumes which would have been used had the patients been treated preoperatively. Hypothetical preoperative treatment plans were generated for each patient based on these modified contours. The hypothetical preoperative treatment plans were then compared to the actual postoperative plans with respect to dose-volume parameters including lung mean dose, lung V20, heart V20 and V30, and mean doses to abdominal structures. Target volumes were smaller with preoperative treatment, with an average reduction of 23%. Comparative dose-volume histogram (DVH) analysis showed the resultant composite lung doses were reduced in the preoperative plans by 50-79%. In all patients, the proportion of lungs receiving at least 20 Gy (V20) was substantially reduced using preoperative treatment (1.9% vs. 9.7% in the 3-D conformal patient; mean of 3.1% vs. 17.6% in the intensity modulated radiation therapy patients). Likewise, the volume of heart receiving at least 30 Gy was dramatically reduced in all preoperative plans (15.8% vs. 35.4%). Doses to the kidneys, liver and spinal cord were comparable in both approaches. Preoperative treatment of GE junction and proximal gastric cancer patients offers the potential to decrease the radiation dose received by normal thoracic structures.

Initial evaluation of 18F-fluorothymidine (FLT) PET/CT scanning for primary pancreatic cancer.

PURPOSE: The aim of this study was to evaluate the potential of (18)F-fluorothymidine (FLT) PET/CT for imaging pancreatic adenocarcinoma. METHODS: This was a pilot study of five patients (four males, one female) with newly diagnosed and previously untreated pancreatic adenocarcinoma. Patients underwent FLT PET/CT, (18)F-fluorodeoxyglucose (FDG) PET/CT, and contrast-enhanced CT scanning before treatment. The presence of cancer was confirmed by histopathological analysis at the time of scanning in all five patients. The degree of FLT and FDG uptake at the primary tumor site was assessed using visual interpretation and semi-quantitative SUV analyses. RESULTS: The primary tumor size ranged from 2.5 x 2.8 cm to 3.5 x 7.0 cm. The SUV of FLT uptake within the primary tumor ranged from 2.1 to 3.1. Using visual interpretation, the primary cancer could be detected from background activity in two of five patients (40%) on FLT PET/CT. By comparison, FDG uptake was higher in each patient with a SUV range of 3.4 to 10.8, and the primary cancer could be detected from background in all five patients (100%). CONCLUSIONS: In this pilot study of five patients with primary pancreatic adenocarcinoma, FLT PET/CT scanning showed poor lesion detectability and relatively low levels of radiotracer uptake in the primary tumor.

Pancreatic tumors show high levels of hypoxia.

PURPOSE: Because of the dismal outcomes of conventional therapies for pancreatic carcinomas, we postulated that hypoxia may exist within these tumors. METHODS AND MATERIALS: Seven sequential patients with adenocarcinomas of the pancreas consented to intraoperative measurements of tumor oxygenation using the Eppendorf (Hamburg, Germany) polargraphic electrode. RESULTS: All 7 tumors demonstrated significant tumor hypoxia. In contrast, adjacent normal pancreas showed normal oxygenation. CONCLUSION: Tumor hypoxia exists within pancreatic cancers.

Candidate genes for the hypoxic tumor phenotype.

In this study, we have analyzed changes induced by hypoxia at the transcriptional level of genes that could be responsible for a more aggressive phenotype. Using a series of DNA array membranes, we identified a group of hypoxia-induced genes that included plasminogen activator inhibitor-1 (PAI-1), insulin-like growth factor-binding protein 3 (IGFBP-3), endothelin-2, low-density lipoprotein receptor-related protein (LRP), BCL2-interacting killer (BIK), migration-inhibitory factor (MIF), matrix metalloproteinase-13 (MMP-13), fibroblast growth factor-3 (FGF-3), GADD45, and vascular endothelial growth factor (VEGF). The induction of each gene was confirmed by Northern blot analysis in two different squamous cell carcinoma-derived cell lines. We also analyzed the kinetics of PAI-1 induction by hypoxia in more detail because it is a secreted protein that may serve as a useful molecular marker of hypoxia. On exposure to hypoxia, there was a gradual increase in PAI-1 mRNA between 2 and 24 h of hypoxia followed by a rapid decay after 2 h of reoxygenation. PAI-1 levels were also measured in the serum of a small group of head and neck cancer patients and were found to correlate with the degree of tumor hypoxia found in these patients.

Hypoxic activation of nuclear factor-kappa B is mediated by a Ras and Raf signaling pathway and does not involve MAP kinase (ERK1 or ERK2).

We have previously shown that hypoxia causes the activation of nuclear factor-kappa B (NF-kappa B), and the phosphorylation of its inhibitory subunit, I kappa B alpha, on tyrosine residues. With the use of dominant negative mutants of Ha-Ras and Raf-1, we investigated some of the early signaling events leading to the activation of NF-kappa B by hypoxia. Both dominant negative alleles of Ha-Ras and Raf-1 inhibited NF-kappa B induction by hypoxia, suggesting that the hypoxia-induced pathway of NF-kappa B induction is dependent on Ras and Raf-1 kinase activity. Furthermore, although conditions of low oxygen can also activate mitogen-activated protein kinases (ERK1 and ERK2), these kinases do not appear to be involved in regulating NF-kappa B by low oxygen conditions, as dominant negative mutants of mitogen-activated protein kinase do not inhibit NF-kappa B activation by hypoxia. Since Ras and Raf-1 have been previously shown to work downstream from membrane-associated tyrosine kinases such as Src, we determined if the Src membrane-associated kinase was also activated by low oxygen conditions. We detected an increase in Src proto-oncogene activity within 15-30 min of cellular exposure to hypoxia. We postulate that Src activation by hypoxia may be one of the earliest events that precedes Ras activation in the signaling cascade which ultimately leads to the phosphorylation and dissociation of the inhibitory subunit of NF-kappa B, I kappa B alpha.

Modulation of potassium channels by protein tyrosine kinase inhibitors.

Exposure of non-excitatory cells to the tyrosine kinase (PTK) inhibitors, genistein, herbimycin A, and tyrphostin, induced at least two families of K+ currents. The first, a TEA-insensitive slow-inactivating K+ current, is induced within 3 min following treatment with 140 mM genistein or 100 nM herbimycin A. The second current, a TEA-sensitive delayed rectifier, is induced within 30 min following treatment with 50 mM genistein or 10 nM herbimycin A. Currents with similar biophysical and pharmacological characteristics are induced in these cells following exposure to ionizing radiation. The radiation-induced currents are inhibited by pretreatment with the free radical scavenger, N-Acetyl L-Cysteine, or by pretreatment with the protein kinase C inhibitor, staurosporine; those induced by PTK inhibitors are not. The latter, therefore, do not appear to be mediated through free radicals or require serine/threonine phosphorylation for activation. Once the channels are activated by the PTK inhibitors, phosphorylation of the channel at serine/threonine residues results in slower inactivation of the induced current. We propose that protein tyrosine phosphorylation of the K+ channel protein itself or of a factor that interacts with it maintains the K+ channels of non-excitatory cells in a closed state. Following exposure to ionizing radiation, free radical-induced activation of serine/threonine kinase(s) results in phosphorylation of the channel and/or inactivation of a tyrosine kinase that in turn leads to activation of the K+ channels.

Analysis of HSF-1 phosphorylation in A549 cells treated with a variety of stresses.

In the absence of stress, heat shock transcription factor-1 (HSF-1) exists as a monomer. After the treatment of cells with variety of stresses, HSF-1 forms a trimer and binds to the heat shock element (HSE), a motif consisting of three consecutive NGAAN sequences located in an inverted orientation upstream of the heat shock genes. HSF-1 is then phosphorylated causing transactivation of heat shock mRNAs. Treatment of cells with some of the stresses has been shown to increase HSF binding to HSE without detectably increasing the synthesis of heat shock mRNAs. Here we used antibody specific to HSF-1 to detect its phosphorylation status following exposure of A549, a human lung carcinoma cell line to a variety of stresses in order to correlate HSF-1 phosphorylation with its transactivation ability. Our studies show that HSF-1 is phosphorylated following heat shock (43 degrees C for 1 h), hypoxia (5 h exposure to 0.02% oxygen), 8% ethanol (1 h exposure at 37 degrees C), or 200 microM sodium arsenite (1 h exposure at 37 degrees C). All such stresses have previously been shown to increase the synthesis of heat shock proteins (hsps). However, there are no detectable increases in HSF-1 phosphorylation after the treatment of cells with X-irradiation (2-8 Gy) or 100 microM canavanine, an amino acid analogue (1 h exposure at 37 degrees C). Treatment of cells with X-irradiation increases HSF binding to HSE without increasing the synthesis of hsps, while treatment of cells with canavanine has been shown to increase the synthesis of hsps.(ABSTRACT TRUNCATED AT 250 WORDS)

Activators of protein kinase C selectively mediate cellular cytotoxicity to hypoxic cells and not aerobic cells.

PURPOSE: By understanding the signal transduction pathways through which a cell responds to changes in environmental oxygen levels, we may be able to therapeutically exploit this response by manipulating these pathways. MATERIALS AND METHODS: The human adenocarcinoma cell line A549 was exposed to varying durations of hypoxia alone and then plated for survival, or treated with PKC activating agents for 1 h before plating for survival. Western blots were used to determine the kinetics of PKC epsilon and phospholipase C induction. RESULTS: The level of hypoxic killing was directly related to the time of exposure and inversely related to the level of oxygen in the environment. Exposure of the cells to protein kinase C (PKC) activators for 1 h after chronic hypoxic exposure increased cell killing by at least an additional three logs beyond that found for hypoxia alone. Treatment of cells with an inactive phorbol ester 4 alpha-phorbol-12,13-didecanoate (PDA) resulted in no increase in hypoxic cell killing, even at the highest concentrations of PDA which produced no detectable toxicity under normal aerobic conditions. Using inhibitors of phospholipases A2 and C, we were able to completely inhibit the additional hypoxic cell killing induced by TPA, but not the uninduced hypoxic cell killing. CONCLUSION: These studies suggest that accumulation of phospholipid breakdown products may be responsible for TPA induced cell killing, and that hypoxic cells differ from aerobic cells in their ability to tolerate these products.

The regulation of GRP78 and messenger RNA levels by hypoxia is modulated by protein kinase C activators and inhibitors.

In this study, we have shown that steady-state levels of glucose-regulated 78 kDa (GRP78) protein and messenger RNA increase during a 5-h exposure to 0.02% oxygen. This increase in GRP78 protein and mRNA induced by hypoxia can be abolished by a 1-h pretreatment of cells before hypoxia with the protein kinase C (PKC) inhibitors staurosporine and H7 at concentrations at which the drugs themselves do not cause cytotoxicity. Although all studies using protein kinase inhibitors must be interpreted with caution, staurosporine and H7 have been shown to be potent inhibitors of PKC activity, suggesting a role for PKC in mediating the transcriptional regulation of GRP78 by hypoxia. Further support for PKC in regulating GRP78 gene expression by hypoxia stems from gel-mobility shift studies in mixtures of nuclear extracts from aerobic or hypoxic cells with a 36 bp region of the GRP78 promoter (-170 to -135). Binding of this factor could be inhibited by pretreating cells with the PKC inhibitor staurosporine before hypoxia or activated by treating cells with the PKC-activating phorbol ester TPA. These data suggest that activation of this hypoxia-responsive factor is sensitive to oxygen levels and seems to be mediated through a PKC signal transduction pathway.

Hypoxia causes the activation of nuclear factor kappa B through the phosphorylation of I kappa B alpha on tyrosine residues.

The response of mammalian cells to stress is controlled by transcriptional regulatory proteins such as nuclear factor kappa B (NF-kappa B) to induce a wide variety of early response genes. In this report, we show that exposure of cells to hypoxia (0.02% O2) results in I kappa B alpha degradation, increased NF-kappa B DNA binding activity, and transactivation of a reporter gene construct containing two NF-kappa B DNA binding sites. Pretreatment of cells with protein tyrosine kinase inhibitors and the dominant negative allele of c-Raf-1 (Raf 301) inhibited I kappa B alpha degradation, NF-kappa B binding, and transactivation of kappa B reporter constructs by hypoxia. To demonstrate a direct link between changes in the phosphorylation pattern of I kappa B alpha with NF-kappa B activation, we immunoprecipitated I kappa B alpha after varying times of hypoxic exposure and found that its tyrosine phosphorylation status increased during hypoxic exposure. Inhibition of the transfer of tyrosine phosphoryl groups onto I kappa B alpha prevented I kappa B alpha degradation and NF-kappa B binding. In comparison to other activators of NF-kappa B such as phorbol myristate acetate or tumor necrosis factor, we did not detect changes in the tyrosine phosphorylation status of I kappa B alpha following treatment with either of these agents. These results suggest that tyrosine phosphorylation of I kappa B alpha during hypoxia is an important proximal step which precedes its dissociation and degradation from NF-kappa B.

Increased cytotoxicity of chronic hypoxic cells by molecular inhibition of GRP78 induction.

PURPOSE: To develop a molecular strategy of increasing cytotoxicity of chronically hypoxic cells by inhibiting Glucose Regulated Protein 78 kDal (GRP78) induction. METHODS AND MATERIALS: A mutant nonGRP78 inducing cell line (78WO) was developed from its parent (DG44) by overexpressing antisense GRP78 mRNA. Following exposure to varying durations of hypoxia, Northern and Western blot analysis were used to characterize the amount of GRP78 expression both at the RNA and protein level. Hypoxia was achieved by placing cells in specially designed hypoxic chambers which were subjected to successive rounds of evacuation and flushing with 95% CO2/N2 to reduce the oxygen in the environment to 0.02% oxygen. After treatment with hypoxia, cells were assayed for colony forming ability. RESULTS: GRP78 mRNA and protein induction following exposure to hypoxia was 3-4 fold lower in the 78WO cell line than in the parental DG44 cell line. Furthermore, it was observed that there was no difference in the cytotoxicity of 78WO and DG44 cells after 10 h of hypoxia. However, after 15 h of hypoxia, the survival of 78WO cells decreased by 1 log and after 20 h of hypoxia, the survival of 78WO decreased by another log. CONCLUSION: These results show that stress protein induction is important for cellular survival to chronic hypoxia and that inhibition of GRP78 induction may represent a novel therapeutic strategy by selectively sensitizing chronically hypoxic cells within solid tumors.

Activation of potassium channels by hypoxia and reoxygenation in the human lung adenocarcinoma cell line A549.

Active oxygen species are generated in cells during pathophysiologic conditions such as inflammation and postischemic reperfusion. If oxygen radical scavengers are added before reperfusion, then the magnitude of injury is reduced. We investigated whether free radicals generated following exposure to hypoxia and reoxygenation activate voltage-dependent K+ ion channels in tumor cells in vitro. Using the technique of whole cell voltage clamping, we recorded currents from two families of potassium (K+) channels that were activated following reoxygenation. One of these groups possessed the electrophysical characteristics of a tetraethylammonium (TEA)-sensitive delayed rectifier channel and the other possessed characteristics of a Tea-insensitive slow inactivating channel. We present evidence which suggests that K+ channels are activated following reoxygenation but not during the hypoxia phase. The K+ currents decayed with time following reoxygenation. The decay characteristics of the K+ currents depended on the duration and level of hypoxia to which the cells were exposed. To determine whether activation of K+ channels by reoxygenation was initiated by free radicals, we pretreated cells with N-Acetyl L-Cysteine (NAC), a free radical scavenger, and found that this pretreatment abolished the currents induced by reoxygenation. We also present evidence that free radicals do not directly act on the channel itself, but activate a protein kinase which, in turn, activates the K+ channels. Taken together, these results indicate that one of the early responses to oxidative stress is the activation of K+ currents.

Potassium-channel activation in response to low doses of gamma-irradiation involves reactive oxygen intermediates in nonexcitatory cells.

Active oxygen species are generated during pathophysiologic conditions such as inflammation and ionizing radiation exposure. We tested the hypothesis that an early cellular event in response to these species involves regulation of ion channels. We exposed cells to gamma-irradiation or treated them with hydrogen peroxide, xanthine/xanthine oxidase, or [3H]thymidine and then monitored channel activity by the technique of whole-cell voltage clamping. Recordings showed that both normal and tumor cells exhibit an increase in K+ currents after treatment with radiation, H2O2, and xanthine/xanthine oxidase but not with high specific activity [3H]thymidine, suggesting that the signal for K+ channel activation originates at the cell membrane. A single noncytotoxic dose of 10 cGy induced measurable levels of K+ currents, suggesting that the induction of currents regulates biochemical changes in response to stress. To test whether channel activity is sensitive to active oxygen species, we pretreated cells with N-acetyl-L-cysteine (NAC) to increase cellular pools of free radical scavengers before radiation. In NAC-pretreated cells, K+ channel activation by gamma-irradiation was abolished. It has previously been shown that protein kinase C (PKC) is activated by ionizing radiation and can regulate K+ channels in some cells. However, the effect of radiation on induction of K+ channel activity was independent of PKC, since cells chronically exposed to phorbol esters still produced K+ currents after radiation. These results suggest that an early cellular response to oxidative stress is the activation of K+ channels.

The influence of chronic anaemia on the radiosensitivity of two mouse tumours.

There is clear clinical evidence that tumours in anaemic patients are difficult to control with radiotherapy. We have studied the radiosensitivity of two transplantable mouse tumours, the SCCVII/St carcinoma and the KHT sarcoma in hosts made anaemic either with an iron poor diet or as a result of tumour growth. The haemoglobin level and haematocrits of mice on the low iron diet fell to about 60% of normal within 11 weeks. The number of clonogenic cells after a single X-ray dose of 20 Gy was slightly lower (P less than 0.05) in the anaemic animals (2.3 X 10(4) g-1) than in controls (5.2 X 10(4) g-1) though there was no significant difference in the surviving fractions. Mice bearing KHT tumours became anaemic with haematocrits falling to 65% of normal as their tumours grew from 300-1200 mg. A second 'test' tumour was implanted one week after the first 'anaemia-inducing' tumour so that estimates of radiosensitivity could all be carried out on tumours within the same size range (150-300 mg). Radiosensitivity was significantly greater in the most anaemic hosts with 2.2 X 10(4) cells g-1 surviving a dose of 20 Gy compared with 6.7 x 10(4) g-1 in controls (P less than 0.01). These results are consistent with most published data for mouse tumours though not for many human tumours.