A Phase 1 Study of a CDH6-Targeting Antibody-Drug Conjugate in Patients with Advanced Solid Tumors with Evaluation of Inflammatory and Neurological Adverse Events.

PURPOSE: This first-in-human study (NCT02947152) evaluated the safety, tolerability, pharmacokinetics, and preliminary efficacy of HKT288, a first-in-class CDH6-targeting antibody-drug conjugate (ADC). EXPERIMENTAL DESIGN: HKT288 was administered intravenously (IV) every 3 weeks until patients experienced unacceptable toxicity or progressive disease (PD). The starting dose of 0.3 mg/kg was determined based on the highest nonseverely toxic dose in monkeys, which was 2 mg/kg IV weekly. Based on preclinical toxicology, skin, eyes, bone marrow, and liver were expected targets of toxicity. RESULTS: Nine patients were enrolled: 5 with renal cell carcinoma and 4 with epithelial ovarian cancer. The best overall response on the 0.3 mg/kg cohort in patients with measurable disease was RECIST v1.1 stable disease in 3 patients and PD in 2 patients. The most frequent adverse events (AEs) regardless of causality were pyrexia (44.4%), constipation (44.4%), fatigue (33.3%), and vomiting (33.3%). Three suspected-related neurologic AEs (Grade 2) were reported on the 0.75 mg/kg cohort: seizure in 1 patient and another patient with aphasia and encephalopathy. Further studies were unable to identify the underlying mechanism of the neurologic AEs, and the study was terminated early. CONCLUSIONS: Preclinical toxicology did not predict the neurotoxicity observed with HKT288, and a comprehensive assessment performed post hoc did not identify the mechanism of toxicity. The development of further CDH6-targeting ADCs should be pursued with caution.

Increased Mortality and Comorbidity Associated With Leber's Hereditary Optic Neuropathy: A Nationwide Cohort Study.

Purpose: Leber's hereditary optic neuropathy (LHON) is a mitochondrial genetic disease in which optic neuropathy is considered a key feature. Several other manifestations of LHON have been reported; however, only little is known of their incidence and the life expectancy in LHON patients. Methods: This study, based on Danish nationwide health registries, included 141 patients diagnosed with LHON and 297 unaffected family members in the maternal line. The incidence of comorbidities and mortality for patients with LHON and unaffected family members was compared with that in the general population. Results: Having LHON was associated with an almost 2-fold risk of mortality with a rate ratio (RR) of 1.95 (95% confidence interval [CI]: 1.47-2.59; P < 0.001). The incidence of several diseases was increased for LHON patients, but not for family members. The incidence of stroke was 5.73 per 1000 patient-years for LHON patients compared to 2.33 for the general population, and the RR was 2.38 (95% CI: 1.58-3.58; P < 0.001). The incidence of demyelinating disorders was 2.24 compared to 0.21 for the general population; RR was 12.89 (95% CI: 6.70-24.77; P < 0.001). A 4-fold risk of dementia was seen for LHON patients (RR: 4.26, 95% CI: 1.91-9.48; P < 0.001), incidence 1.45 for LHON and 0.37 for the general population. Moreover, LHON patients had an increased risk of epilepsy, atherosclerosis, nerve symptoms, neuropathy, and alcohol-related disorders. Conclusions: The manifestation of LHON was associated with increased mortality and increased incidence of several disorders including stroke, demyelinating disorder, dementia, and epilepsy.

Discovery and Optimization of HKT288, a Cadherin-6-Targeting ADC for the Treatment of Ovarian and Renal Cancers.

Despite an improving therapeutic landscape, significant challenges remain in treating the majority of patients with advanced ovarian or renal cancer. We identified the cell-cell adhesion molecule cadherin-6 (CDH6) as a lineage gene having significant differential expression in ovarian and kidney cancers. HKT288 is an optimized CDH6-targeting DM4-based antibody-drug conjugate (ADC) developed for the treatment of these diseases. Our study provides mechanistic evidence supporting the importance of linker choice for optimal antitumor activity and highlights CDH6 as an antigen for biotherapeutic development. To more robustly predict patient benefit of targeting CDH6, we incorporate a population-based patient-derived xenograft (PDX) clinical trial (PCT) to capture the heterogeneity of response across an unselected cohort of 30 models-a novel preclinical approach in ADC development. HKT288 induces durable tumor regressions of ovarian and renal cancer models in vivo, including 40% of models on the PCT, and features a preclinical safety profile supportive of progression toward clinical evaluation.Significance: We identify CDH6 as a target for biotherapeutics development and demonstrate how an integrated pharmacology strategy that incorporates mechanistic pharmacodynamics and toxicology studies provides a rich dataset for optimizing the therapeutic format. We highlight how a population-based PDX clinical trial and retrospective biomarker analysis can provide correlates of activity and response to guide initial patient selection for first-in-human trials of HKT288. Cancer Discov; 7(9); 1030-45. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 920.

Persistent effects of Libby amphibole and amosite asbestos following subchronic inhalation in rats.

BACKGROUND: Human exposure to Libby amphibole (LA) asbestos increases risk of lung cancer, mesothelioma, and non-malignant respiratory disease. This study evaluated potency and time-course effects of LA and positive control amosite (AM) asbestos fibers in male F344 rats following nose-only inhalation exposure. METHODS: Rats were exposed to air, LA (0.5, 3.5, or 25.0 mg/m(3) targets), or AM (3.5 mg/m(3) target) for 10 days and assessed for markers of lung inflammation, injury, and cell proliferation. Short-term results guided concentration levels for a stop-exposure study in which rats were exposed to air, LA (1.0, 3.3, or 10.0 mg/m(3)), or AM (3.3 mg/m(3)) 6 h/day, 5 days/week for 13 weeks, and assessed 1 day, 1, 3, and 18 months post-exposure. Fibers were relatively short; for 10 mg/m(3) LA, mean length of all structures was 3.7 µm and 1% were longer than 20 µm. RESULTS: Ten days exposure to 25.0 mg/m(3) LA resulted in significantly increased lung inflammation, fibrosis, bronchiolar epithelial cell proliferation and hyperplasia, and inflammatory cytokine gene expression compared to air. Exposure to 3.5 mg/m(3) LA resulted in modestly higher markers of acute lung injury and inflammation compared to AM. Following 13 weeks exposure, lung fiber burdens correlated with exposure mass concentrations, declining gradually over 18 months. LA (3.3 and 10.0 mg/m(3)) and AM produced significantly higher bronchoalveolar lavage markers of inflammation and lung tissue cytokines, Akt, and MAPK/ERK pathway components compared to air control from 1 day to 3 months post-exposure. Histopathology showed alveolar inflammation and interstitial fibrosis in all fiber-exposed groups up to 18 months post-exposure. Positive dose trends for incidence of alveolar epithelial hyperplasia and bronchiolar/alveolar adenoma or carcinoma were observed among LA groups. CONCLUSIONS: Inhalation of relatively short LA fibers produced inflammatory, fibrogenic, and tumorigenic effects in rats which replicate essential attributes of asbestos-related disease in exposed humans. Fiber burden, inflammation, and activation of growth factor pathways may persist and contribute to lung tumorigenesis long after initial LA exposure. Fiber burden data are being used to develop a dosimetry model for LA fibers, which may provide insights on mode of action for hazard assessment.

Pten loss promotes MAPK pathway dependency in HER2/neu breast carcinomas.

Loss of the tumor suppressor gene PTEN is implicated in breast cancer progression and resistance to targeted therapies, and is thought to promote tumorigenesis by activating PI3K signaling. In a transgenic model of breast cancer, Pten suppression using a tetracycline-regulatable short hairpin (sh)RNA cooperates with human epidermal growth factor receptor 2 (HER2/neu), leading to aggressive and metastatic disease with elevated signaling through PI3K and, surprisingly, the mitogen-activated protein kinase (MAPK) pathway. Restoring Pten function is sufficient to down-regulate both PI3K and MAPK signaling and triggers dramatic tumor regression. Pharmacologic inhibition of MAPK signaling produces similar effects to Pten restoration, suggesting that the MAPK pathway contributes to the maintenance of advanced breast cancers harboring Pten loss.

Late radiation sequelae as a consequence of breast-conserving therapy with cobalt irradiation aggravated by various risk factors.

This report deals with a 71-year-old female patient who developed cancer in her right breast 20 years ago, underwent breast-conserving surgery and received normofractionated radiotherapy with a 60Co unit. 19 years later, fibroids and calcified tissue appeared in her right mammary fold. Furthermore, a deep ulceration developed in this region during chemotherapy of bronchial carcinoma. Apart from being a Type 2 diabetic with arterial hypertension, she was also a habitual smoker. After extensive wound debridement and vacuum-assisted sealing therapy, the affected ribs were dissected and a latissimus dorsi flap was implanted. Our focus here is on the interaction of contributing risks for the development of late radiation sequelae, such as physical (especially unintended hot spots during 60Co irradiation) and pathophysiological factors (comorbidities and morbid affections). Fortunately, side-effects such as these are rare nowadays. As this case shows, however, they can be effectively handled by employing modern plastic surgery techniques.

An antibody that locks HER3 in the inactive conformation inhibits tumor growth driven by HER2 or neuregulin.

HER2/HER3 dimerization resulting from overexpression of HER2 or neuregulin (NRG1) in cancer leads to HER3-mediated oncogenic activation of phosphoinositide 3-kinase (PI3K) signaling. Although ligand-blocking HER3 antibodies inhibit NRG1-driven tumor growth, they are ineffective against HER2-driven tumor growth because HER2 activates HER3 in a ligand-independent manner. In this study, we describe a novel HER3 monoclonal antibody (LJM716) that can neutralize multiple modes of HER3 activation, making it a superior candidate for clinical translation as a therapeutic candidate. LJM716 was a potent inhibitor of HER3/AKT phosphorylation and proliferation in HER2-amplified and NRG1-expressing cancer cells, and it displayed single-agent efficacy in tumor xenograft models. Combining LJM716 with agents that target HER2 or EGFR produced synergistic antitumor activity in vitro and in vivo. In particular, combining LJM716 with trastuzumab produced a more potent inhibition of signaling and cell proliferation than trastuzumab/pertuzumab combinations with similar activity in vivo. To elucidate its mechanism of action, we solved the structure of LJM716 bound to HER3, finding that LJM716 bound to an epitope, within domains 2 and 4, that traps HER3 in an inactive conformation. Taken together, our findings establish that LJM716 possesses a novel mechanism of action that, in combination with HER2- or EGFR-targeted agents, may leverage their clinical efficacy in ErbB-driven cancers.

Neuroprotection by oxygen in acute transient focal cerebral ischemia is dose dependent and shows superiority of hyperbaric oxygenation.

The neuroprotective effect of oxygen after acute stroke in rats has been shown previously. However, the question of optimal dosing still remains unanswered. Thus, we investigated the use of oxygen at different concentrations by either normobaric oxygenation (NBO) or hyperbaric oxygenation (HBO) at different pressures in a model of transient ischemia/reperfusion in rats. Animals underwent 90 min of middle cerebral artery occlusion (MCAO) followed by 90 min of reperfusion before oxygen treatment. Oxygen was applied either by NBO (100% O(2); 1.0 absolute atmosphere, ATA) or HBO (100% O(2); 1.5, 2.0, 2.5 or 3.0 ATA) for 1 h. Primary endpoints were infarct volume and clinical outcome measured 24 h and 7 days following the MCAO. A statistically significant and long-lasting reduction in infarct volume was seen in the HBO 2.5 ATA and 3.0 ATA groups over a period of 7 days. The reduced infarct volume was accompanied with a statistically significant improvement in clinical outcome in the high-dose oxygen-treated groups. The presented data indicate that oxygen is a highly neuroprotective molecule in transient focal cerebral ischemia in rats, when applied early and at high doses. The effect is dose dependent and shows a superiority of HBO over NBO, when the primary endpoints infarct volume reduction and clinical outcome are analyzed. These data are important for the development of new acute stroke treatment studies in humans.

p32 is a novel mammalian Lgl binding protein that enhances the activity of protein kinase Czeta and regulates cell polarity.

Lgl (lethal giant larvae) plays an important role in cell polarity. Atypical protein kinase C (aPKC) binds to and phosphorylates Lgl, and the phosphorylation negatively regulates Lgl activity. In this study, we identify p32 as a novel Lgl binding protein that directly binds to a domain on mammalian Lgl2 (mLgl2), which contains the aPKC phosphorylation site. p32 also binds to PKCzeta, and the three proteins form a transient ternary complex. When p32 is bound, PKCzeta is stimulated to phosphorylate mLgl2 more efficiently. p32 overexpression in Madin-Darby canine kidney cells cultured in a 3D matrix induces an expansion of the actin-enriched apical membrane domain and disrupts cell polarity. Addition of PKCzeta inhibitor blocks apical actin accumulation, which is rescued by p32 overexpression. p32 knockdown by short hairpin RNA also induces cell polarity defects. Collectively, our data indicate that p32 is a novel regulator of cell polarity that forms a complex with mLgl2 and aPKC and enhances aPKC activity.

Tissue manganese concentrations in young male rhesus monkeys following subchronic manganese sulfate inhalation.

High-dose human exposure to manganese results in manganese accumulation in the basal ganglia and dopaminergic neuropathology. Occupational manganese neurotoxicity is most frequently linked with manganese oxide inhalation; however, exposure to other forms of manganese may lead to higher body burdens. The objective of this study was to determine tissue manganese concentrations in rhesus monkeys following subchronic (6 h/day, 5 days/week) manganese sulfate (MnSO(4)) inhalation. A group of monkeys were exposed to either air or MnSO(4) (0.06, 0.3, or 1.5 mg Mn/m(3)) for 65 exposure days before tissue analysis. Additional monkeys were exposed to MnSO(4) at 1.5 mg Mn/m(3) for 15 or 33 exposure days and evaluated immediately thereafter or for 65 exposure days followed by a 45- or 90-day delay before evaluation. Tissue manganese concentrations depended upon the aerosol concentration, exposure duration, and tissue. Monkeys exposed to MnSO(4) at > or = 0.06 mg Mn/m(3) for 65 exposure days or to MnSO(4) at 1.5 mg Mn/m(3) for > or = 15 exposure days developed increased manganese concentrations in the olfactory epithelium, olfactory bulb, olfactory cortex, globus pallidus, putamen, and cerebellum. The olfactory epithelium, olfactory bulb, globus pallidus, caudate, putamen, pituitary gland, and bile developed the greatest relative increase in manganese concentration following MnSO(4) exposure. Tissue manganese concentrations returned to levels observed in the air-exposed animals by 90 days after the end of the subchronic MnSO(4) exposure. These results provide an improved understanding of MnSO(4) exposure conditions that lead to increased concentrations of manganese within the nonhuman primate brain and other tissues.

Respiration in Sprague-Dawley rats during pregnancy.

Minute ventilation and tidal volume increase in humans during pregnancy. Little data exists, however, on the respiration in pregnant rats, despite their widespread use as an animal model. Since respiration will affect the pharmacokinetics of volatile compounds and ultimately the dose to the fetus, we conducted a study to evaluate respiration in rats during pregnancy. Whole-body plethysmography was used to measure the breathing frequency and tidal volume approximately every other day from gestation day (GD) 1 to 21 in 16 timed pregnant and 16 nonpregnant, female, Sprague-Dawley rats. Minute ventilation was calculated as a product of the breathing frequency and tidal volume, and the body weight of each rat was used to determine the scaled ventilation. Multivariate analysis of variance methods for a repeated-measures design were used to analyze the respiratory data. Breathing frequency was not affected by pregnancy; however, tidal volume was somewhat greater in pregnant versus nonpregnant rats. The increase in tidal volume resulted in significantly increased minute ventilation in pregnant rats compared to nonpregnant rats during the latter period of gestation. Due to the increased body weight of the pregnant rats, the scaled ventilation at the end of gestation was significantly lower in pregnant rats compared to nonpregnant rats. This study provides important reference values that can be used in pharmacokinetic models during pregnancy.

Maternal-fetal distribution of manganese in the rat following inhalation exposure to manganese sulfate.

Studies examining the pharmacokinetics of manganese during pregnancy have largely focused on the oral route of exposure and have shown that the amount of manganese that crosses the rodent placenta is low. However, limited information exists regarding the distribution of manganese in fetal tissues following inhalation. The objective of this study was to determine manganese body burden in CD rats and fetuses following inhalation of a MnSO4 aerosol during pregnancy. Animals were evaluated following pre-breeding (2 weeks), mating (up to 14 days) and gestational (from gestation day (GD) 0 though 20) exposure to air or MnSO4 (0.05, 0.5, or 1 mg Mn/m(3)) for 6h/day, 7 days/week. The following maternal samples were collected for manganese analysis: whole blood, lung, pancreas, liver, brain, femur, and placenta. Fetal tissues were examined on GD 20 and included whole blood, lung, liver, brain, and skull cap. Maternal lung manganese concentrations were increased following exposure to MnSO4 at >or=0.05 mg Mn/m(3). Maternal brain and placenta manganese concentrations were increased following exposure of pregnant rats to MnSO4 at >or=0.5 mg Mn/m(3). Increased fetal liver manganese concentrations were observed following in utero exposure to MnSO4 at >or=0.5 mg Mn/m(3). Manganese concentrations within all other fetal tissues were not different from air-exposed controls. The results of this study demonstrate that the placenta partially sequesters inhaled manganese, thereby limiting exposure to the fetus.

Tissue manganese concentrations in lactating rats and their offspring following combined in utero and lactation exposure to inhaled manganese sulfate.

There is little information regarding the tissue distribution of manganese in neonates following inhalation. This study determined tissue manganese concentrations in lactating CD rats and their offspring following manganese sulfate (MnSO4) aerosol inhalation. Except for the period of parturition, dams and their offspring were exposed to air or MnSO4 (0.05, 0.5, or 1 mg Mn/m3) for 6 h/day, 7 days/week starting 28 days prior to breeding through postnatal day (PND) 18. Despite increased manganese concentrations in several maternal tissues, MnSO4 inhalation exposure did not affect body weight gain, terminal (PND 18) body weight, or organ weights in the dams. Exposure to MnSO4 at 1 mg Mn/m3 resulted in decreased pup body weights on PND 19 and decreased brain weights in some PND 14 to PND 45 pups. Exposure to MnSO4 at > or =0.05 mg Mn/m3 was associated with increased stomach content, blood, liver, and skull cap manganese concentrations in PND 1 pups, increased brain, lung, and femur manganese concentrations in PND 14 pups, and elevated olfactory bulb, cerebellum, and striatum manganese concentrations in PND 19 pups. When compared to controls, MnSO4 exposure to > or =0.5 mg Mn/m3 increased liver and blood manganese concentrations in PND 14 pups and increased liver, pancreas, and femur manganese concentrations in PND 19 pups. Manganese concentrations returned to control values in all offspring tissues by PND 45 +/- 1. Our data demonstrate that neonatal tissue manganese concentrations observed following MnSO4 inhalation are dependent on the MnSO4 exposure concentration and the age of the animal.

Nasal toxicity of manganese sulfate and manganese phosphate in young male rats following subchronic (13-week) inhalation exposure.

Growing evidence suggests that nasal deposition and transport along the olfactory nerve represents a route by which inhaled manganese and certain other metals are delivered to the rodent brain. The toxicological significance of olfactory transport of manganese remains poorly defined. In rats, repeated intranasal instillation of manganese chloride results in injury to the olfactory epithelium and neurotoxicity as evidenced by increased glial fibrillary acidic protein (GFAP) concentrations in olfactory bulb astrocytes. The purpose of the present study was to further characterize the nasal toxicity of manganese sulfate (MnSO(4)) and manganese phosphate (as hureaulite) in young adult male rats following subchronic (90-day) exposure to air, MnSO(4) (0.01, 0.1, and 0.5 mg Mn/m(3)), or hureaulite (0.1 mg Mn/m(3)). Nasal pathology, brain GFAP levels, and brain manganese concentrations were assessed immediately following the end of the 90-day exposure and 45 days thereafter. Elevated end-of-exposure olfactory bulb, striatum, and cerebellum manganese concentrations were observed following MnSO(4) exposure to > or = 0.01, > or = 0.1, and 0.5 mg Mn/m(3), respectively. Exposure to MnSO(4) or hureaulite did not affect olfactory bulb, cerebellar, or striatal GFAP concentrations. Exposure to MnSO(4) (0.5 mg Mn/m(3)) was also associated with reversible inflammation within the nasal respiratory epithelium, while the olfactory epithelium was unaffected by manganese inhalation. These results confirm that high-dose manganese inhalation can result in nasal toxicity (irritation) and increased delivery of manganese to the brain; however, we could not confirm that manganese inhalation would result in altered brain GFAP concentrations.

Influence of antioxidants on the sun protection properties of hair care products.

Aim of this study was to determine the influence of commercially available antioxidants on sun protection properties of hair care products. To evaluate changes of human hair L*a*b-color measurements, tensile strength measurements and high pressure dynamic scanning calorimetry (HPDSC) measurements were carried out. To have a measure for the concentration of the activity of the reactive species, causing hair or color damage, chemiluminescence measurements were carried out. Before the test with the antioxidants experiments were carried out to evaluate effects of varied artificial weathering conditions on physical properties of hair. Here high relative humidity (85%) and low radiant flux (600W/m2) exhibited the biggest changes in natural hair color but the lowest changes in the in tensile strength and HPDSC measurements. All of the tested antioxidants reduced the chemiluminescence level when used in a pre-sun or after-sun formulation. According to the HPDSC measurements the antioxidants showed a slight increase of the peak temperature and therewith a hint towards a protection effect when used in a pre-sun or after-sun product. In contrast thereto some of the antioxidants reduced the tensile strength of sun care products for hair when added. A slight reduction in the lightening of natural hair color could be observed when antioxidants were present in the sun care formulations. The effect of antioxidants in sun care formulations used on dyed hair was strongly dependent on the shade of hair. The addition of some antioxidants yielded significant improvements of the protection properties of the used sun care product in some measurement methods.

Hyperbaric oxygen: does it promote growth or recurrence of malignancy?

It has been a concern that a therapeutic modality recommended as an adjunct to healing and administered to promote proliferation of fibroblasts, epithelial cells and blood vessels in a wound could also lead to proliferation of malignant cells and angiogenesis in a malignant tumor. The first reported concern that hyperbaric oxygen (HBO2) might have cancer growth enhancing effects appeared in a paper by Johnson and Lauchlan in 1966. In a series of patients treated with HBO2 radiosensitization, they reported a more frequent than expected incidence of metastases and an unusual pattern of metastases. The published literature from clinical reports, animal studies and cell culture studies are reviewed. Putative mechanisms whereby HBO2 could have carcinogenic effects are discussed. The processes of angiogenesis in wound healing and in cancer growth are compared and contrasted. In vitro, in vivo and clinical studies strongly suggest no more than a neutral effect of HBO2 on tumor growth. In fact some studies suggest a negative impact of HBO2 on malignant progression or formation. For angiogenesis, similarities in wound healing and cancer are striking but significant differences are found including the relative importance of angiogenic factors and the process of cessation of angiogenesis. Tumors that grow in hypoxic environments are more prone to metastases and more lethal to the patient. They are also more likely to mutate toward resistant genotypes. Discussion of postulated mechanisms of carcinogenesis including free radical and immunosuppressive effects points out why they are not likely to enhance or cause cancer growth or initiation. In conclusion, the published literature on tumor angiogenesis mechanisms and other possible mechanisms of cancer causation or accelerated growth provides little basis for HBO2 to enhance malignant growth or metastases. A history of malignancy should not be considered a contraindication for HBO2 therapy.

Heterotopic calcification as a late radiation effect: report of 15 cases.

The LENT-SOMA scoring system for reporting late tissue effects following therapeutic radiation does not include heterotopic calcification as an end-point. Here we report on 15 long-term radiotherapy survivors with significant heterotopic calcifications. In all cases heterotopic calcification was linked to other radiation sequelae, e.g. ulceration, bone necrosis, nerve damage and fibrosis. The median time interval between radiotherapy and the occurrence of heterotopic calcification was 19 years (range 2-31 years). All patients received doses in excess of 40 Gy; overlap of adjacent fields played a role in some cases. It appears that heterotopic calcification can be regarded as end-stage damage following high dose radiotherapy. Heterotopic calcification in conjunction with local tissue breakdown is highly suggestive of previous radiation treatment.

Effects of hyperbaric oxygen and normobaric carbogen on the radiation response of the rat rhabdomyosarcoma R1H.

PURPOSE: Hypoxic tumor cells are an important factor of radioresistance. Hyperbaric oxygen (HBO) and normobaric carbogen (95% oxygen, 5% carbon dioxide) increase the oxygen delivery to tumors. This study was performed to explore changes of tumor oxygenation during a course of fractionated irradiation and to determine the effectiveness of normobaric carbogen and HBO during the final phase of the radiation treatment. METHODS AND MATERIALS: Experiments were performed on the rhabdomyosarcoma R1H growing on WAG/Rij rats. After 20 X-ray fractions of 2 Gy within 4 weeks, oxygen partial pressure (pO2) was measured using the Eppendorf oxygen electrode under ambient conditions, with normobaric carbogen or HBO at a pressure of 240 kPa. Following the 4-week radiation course, a top-up dose of 10-50 Gy was applied in 2-10 fractions of 5 Gy with or without hyperoxygenation. RESULTS: HBO but not carbogen significantly increased the median pO2 in irradiated tumors. The radiation doses to control 50% of tumors were 38.0 Gy, 29.5 Gy, and 25.0 Gy for air, carbogen, and HBO, respectively. Both high oxygen content gas inspirations led to significantly improved tumor responses with oxygen enhancement ratios (OERs) of 1.3 for normobaric carbogen and 1.5 for HBO (air vs. carbogen: p = 0.044; air vs. HBO: p = 0.02; carbogen vs. HBO: p = 0.048). CONCLUSION: Both normobaric carbogen and HBO significantly improved the radiation response of R1H tumors. HBO appeared to be more effective than normobaric carbogen, both with regard to tumor oxygenation and response to irradiation.

The N-terminal domain of Homer/Vesl is a new class II EVH1 domain.

Cellular activities controlled by signal transduction processes such as cell motility and cell growth depend on the tightly regulated assembly of multiprotein complexes. Adapter proteins that specifically interact with their target proteins are key components required for the formation of these assemblies. Ena/VASP-homology 1 (EVH1) domains are small constituents of large modular proteins involved in microfilament assembly that specifically recognize proline-rich regions. EVH1 domain-containing proteins are present in neuronal cells, like the Homer/Vesl protein family that is involved in memory-generating processes. Here, we describe the crystal structure of the murine EVH1 domain of Vesl 2 at 2.2 A resolution. The small globular protein consists of a seven-stranded antiparallel beta-barrel with a C-terminal alpha-helix packing alongside the barrel. A shallow groove running parallel with beta-strand VI forms an extended peptide-binding site. Using peptide library screenings, we present data that demonstrate the high affinity of the Vesl 2 EVH1 domain towards peptide sequences containing a proline-rich core sequence (PPSPF) that requires additional charged amino acid residues on either side for specific binding. Our functional data, substantiated by structural data, demonstrate that the ligand-binding of the Vesl EVH1 domain differs from the interaction characteristics of the previously examined EVH1 domains of the Evl/Mena proteins. Analogous to the Src homology 3 (SH3) domains that bind their cognate ligands in two distinct directions, we therefore propose the existence of two distinct classes of EVH1 domains.

Hyperbaric oxygen therapy for late sequelae in women receiving radiation after breast-conserving surgery.

PURPOSE: Persisting symptomatology after breast-conserving surgery and radiation is frequently reported. In most cases, symptoms in the breast resolve without further treatment. In some instances, however, pain, erythema, and edema can persist for years and can impact the patient's quality of life. Hyperbaric oxygen therapy was shown to be effective as treatment for late radiation sequelae. The objective of this study was to assess the efficacy of hyperbaric oxygen therapy in symptomatic patients after breast cancer treatment. PATIENTS AND METHODS: Forty-four patients with persisting symptomatology after breast-conservation therapy were prospectively observed. Thirty-two women received hyperbaric oxygen therapy in a multiplace chamber for a median of 25 sessions (range, 7-60). One hundred percent oxygen was delivered at 240 kPa for 90-min sessions, 5 times per week. Twelve control patients received no further treatment. Changes throughout the irradiated breast tissue were scored prior to and after hyperbaric oxygen therapy using modified LENT-SOMA criteria. RESULTS: Hyperbaric oxygen therapy patients showed a significant reduction of pain, edema, and erythema scores as compared to untreated controls (p < 0.001). Fibrosis and telangiectasia, however, were not significantly affected by hyperbaric oxygen therapy. Seven of 32 women were free of symptoms after hyperbaric oxygen therapy, whereas all 12 patients in the control group had persisting complaints. CONCLUSIONS: Hyperbaric oxygen therapy should be considered as a treatment option for patients with persisting symptomatology following breast-conserving therapy.