Impact of EGFRA289T/V mutation on relapse pattern in glioblastoma.

BACKGROUND: Molecular factors influence relapse patterns in glioblastoma. The hotspot mutation located at position 289 of the extracellular domain of the epidermal growth factor receptor (EGFRA289mut) is associated with a more infiltrative phenotype. The primary objective of this study was to explore the impact of the EGFRA289 mutation on the pattern of relapse after chemoradiotherapy-based treatment of patients suffering from newly diagnosed glioblastoma. PATIENTS AND METHODS: An ancillary study from a prospective cohort of patients suffering from glioblastoma was conducted. All patients received radiotherapy and concomitant temozolomide. The population was divided into two groups according to EGFRA289 status (mutated versus wild-type). The primary endpoint was the overlap score (varying from 0 to 1) between the initial irradiated tumor volume (Vinit) and the relapse volume (Vr). Secondary endpoints explored the impact of EGFRA289mut on survival. RESULTS: One hundred twenty-eight patients were included and analyzed: 11% had EGFRA289mut glioblastoma (n = 14/128). EGFRA289mut glioblastomas had a relapse pattern that was more marginal than EGFRA289wt glioblastomas: a median overlap score Vinit/Vr of 0.96 was observed in the EGFRA289mut group versus 1 in the EGFRA289wt group (P = 0.05). Half of the population with EGFRA289mut tumor (n = 7/14) had a marginal relapse (i.e. overlap scoreVr/Vinit ≤ 0.95) compared to 23.7% (n = 27/114) in the EGFRA289wt group, P = 0.035. EGFRA289mut did not influence survival. CONCLUSION: We highlighted a link between the EGFRA289 mutation and the relapse pattern in glioblastoma. The independent role of EGFRA289mut and its clinical implication should now be explored in further studies.

[What dose escalation in the treatment of locally advanced non-small cell lung cancer?] / Quelle augmentation de doses dans le traitement des cancers bronchiques non à petites cellules localement avancés ?

Despite significant therapeutic advances in the treatment of locally advanced inoperable non-small cell lung cancer (NSCLC), notably through adjuvant immunotherapy, the rate of therapeutic failure remains high. The use of positron emission tomography with fluorodeoxyglucose (FDG-PET), respiratory motion and intensity modulated radiotherapy (IMRT) have led to therapeutic improvements with reduced toxicity and better local control. The optimal dose to be delivered remains unknown due to discordant results of studies for almost 20 years and the way to define the area to benefit from a dose increase (whole volume, subvolume defined by pre- or per-radiotherapy PET).

Prognostic value of low skeletal muscle mass in patient treated by exclusive curative radiochemotherapy for a NSCLC.

Low skeletal muscle mass is a well-known prognostic factor for patients treated for a non-small-cell lung cancer by surgery or chemotherapy. However, its impact in patients treated by exclusive radiochemotherapy has never been explored. Our study tries to evaluate the prognostic value of low skeletal muscle mass and other antropometric parameters on this population. Clinical, nutritional and anthropometric date were collected for 93 patients treated by radiochemotherapy for a NSCLC. Anthropometric parameters were measured on the PET/CT by two methods. The first method was a manual segmentation at level L3, used to define Muscle Body Area (MBAL3), Visceral Fat Area (VFAL3) and Subcutaneous Fat Area (SCFAL3). The second method was an software (Anthropometer3D), allowing an automatic multislice measurement of Lean Body Mass (LBMAnthro3D), Fat Body Mass (FBMAnthro3D), Muscle Body Mass (MBMAnthro3D), Visceral Fat Mass (VFMAnthro3D), and Sub-Cutaneous Fat Mass (SCFMAnthro3D) on the PET/CT. All anthropometrics parameters were normalised by the patient's height. The primary end point was overall survival time. Univariate and then stepwise multivariate cox analysis were performed for significant parameters. Finally, Spearman's correlation between MBAL3 and MBMAnthro3D was assessed. Forty-one (44%) patients had low skeletal muscle mass. The median overall survival was 18 months for low skeletal muscle mass patients versus 36 months for non-low skeletal muscle mass patients (p = 0.019). Low skeletal muscle mass (HR = 1.806, IC95% [1.09-2.98]), serums albumin level < 35 g/l (HR = 2.203 [1.19-4.09]), Buzby Index < 97.5 (HR = 2.31 [1.23-4.33]), WHO score = 0 (HR = 0.59 [0.31-0.86] and MBMAnthro3D < 8.56 kg/m2 (HR = 2.36 [1.41-3.90]) were the only significant features in univariates analysis. In the stepwise multivariate Cox analysis, only MBMAnthro3D < 8.56 kg/m2 (HR = 2.16, p = 0.003) and WHO score = 0 (HR = 0.59, p = 0.04) were significant. Finally, muscle quantified by MBAL3 and MBMAnthro3D were found to be highly correlated (Spearman = 0.9). Low skeletal muscle mass, assessed on the pre-treatment PET/CT is a powerful prognostic factor in patient treated by radiochemotherapy for a NSCLC. The automatic software Anthropometer3D can easily identify patients a risk that could benefit an adapted therapy.

Benefits of positron emission tomography scans for the evaluation of radiotherapy.

The assessment of tumour response during and after radiotherapy determines the subsequent management of patients (adaptation of treatment plan, monitoring, adjuvant treatment, rescue treatment or palliative care). In addition to its role in extension assessment and therapeutic planning, positron emission tomography combined with computed tomography provides useful functional information for the evaluation of tumour response. The objective of this article is to review published data on positron emission tomography combined with computed tomography as a tool for evaluating external radiotherapy for cancers. Data on positron emission tomography combined with computed tomography scans acquired at different times (during, after initial and after definitive [chemo-]radiotherapy, during post-treatment follow-up) in solid tumours (lung, head and neck, cervix, oesophagus, prostate and rectum) were collected and analysed. Recent recommendations of the National Comprehensive Cancer Network are also reported. Positron emission tomography combined with computed tomography with (18F)-labelled fluorodeoxyglucose has a well-established role in clinical routine after chemoradiotherapy for locally advanced head and neck cancers, particularly to limit the number of neck lymph node dissection. This imaging modality also has a place for the evaluation of initial chemoradiotherapy of oesophageal cancer, including the detection of distant metastases, and for the post-therapeutic evaluation of cervical cancer. Several radiotracers for positron emission tomography combined with computed tomography, such as choline, are also recommended for patients with prostate cancer with biochemical failure. (18F)-fluorodeoxyglucose positron emission tomography combined with computed tomography is optional in many other circumstances and its clinical benefits, possibly in combination with MRI, to assess response to radiotherapy remain a very active area of research.

How to work together between nuclear medicine and radiotherapy departments?

Among the available imaging techniques, functional imaging provided by nuclear medicine departments represents a tool of choice for the oncoradiotherapist for targeting tumour activity, with positron emission tomography as the main modality. Before, during or after radiotherapy, functional imaging helps guide the oncoradiotherapist in making decisions and in the strategic choice of pathology management. Setting up a working group to ensure perfect coordination at all levels is the first step. Key points for a common and coordinated management between the two departments are the definition of an organizational logistic, training of personnel at every levels, standardization of nomenclatures, the choice of adapted and common equipment, implementation of regulatory controls, and research/clinical routine continuum. The availability of functional examinations dedicated to radiotherapy in clinical routine is possible and requires a convergence of teams and a pooling of tools and techniques.

Comparison of Hypermetabolic and Hypoxic Volumes Delineated on [18F]FDG and [18F]Fluoromisonidazole PET/CT in Non-small-cell Lung Cancer Patients.

PURPOSE: The high rates of failure in the radiotherapy target volume suggest that patients with stage II or III non-small-cell lung cancer (NSCLC) should receive an increased total dose of radiotherapy. 2-Deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) and [18F]fluoromisonidazole ([18F]FMISO) (hypoxia) uptake on pre-radiotherapy positron emission tomography (PET)/X-ray computed tomography (CT) have been independently reported to identify intratumor subvolumes at higher risk of relapse after radiotherapy. We have compared the [18F]FDG and [18F]FMISO volumes defined by PET/CT in NSCLC patients included in a prospective study. PROCEDURES: Thirty-four patients with non-resectable lung cancer underwent [18F]FDG and [18F]FMISO PET/CT before (pre-RT) and during radiotherapy (around 42 Gy, per-RT). The criteria were to delineate 40 % and 90 % SUVmax thresholds on [18F]FDG PET/CT (metabolic volumes), and SUV > 1.4 on pre-RT [18F]FMISO PET/CT (hypoxic volume). The functional volumes were delineated within the tumor volume as defined on co-registered CTs. RESULTS: The mean pre-RT and per-RT [18F]FDG volumes were not statistically different (30.4 cc vs 22.2; P = 0.12). The mean pre-RT SUVmax [18F]FDG was higher than per-RT SUVmax (12.7 vs 6.5; P < 0.0001). The mean [18F]FMISO SUVmax and volumes were 2.7 and 1.37 cc, respectively. Volume-based analysis showed good overlap between [18F]FDG and [18F]FMISO for all methods of segmentation but a poor correlation for Jaccard or Dice Indices (DI). The DI maximum was 0.45 for a threshold at 40 or 50 %. CONCLUSION: The correlation between [18F]FDG and [18F]FMISO uptake is low in NSCLC, making it possible to envisage different management strategies as the studies in progress show.

["Definition of target volume: When et how can the radiation oncologist use PET?"] / « Définition des volumes cibles : quand et comment l'oncologue radiothérapeute peut-il utiliser la TEP ? ¼.

PET/CT has become a standard examination in oncology but is probably still underused for radiotherapy planning. However, except for the clinical research data that shows the interest of this examination in considering personalized and adaptive radiotherapy, it is also important in defining target volumes. However, before using it in clinical practice, a few prerequisites are required to know the acquisition and segmentation methods. Ideally, PET/CT should become a standard examination for radiotherapy departments in the same way as planning CT and tomorrow as MRI.

Delineation of lung cancer with FDG PET/CT during radiation therapy.

OBJECTIVES: To propose an easily applicable segmentation method (perPET-RT) for delineation of tumour volume during radiotherapy on interim fluorine 18 fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) in patients with non-small cell lung cancer (NSCLC). MATERIAL AND METHODS: Sixty-seven patients (51 primary tumours, 60 lymph nodes), from 4 prospective studies, underwent an FDG PET/CT scan during the fifth week of radiation therapy, using different generations of PET/CT. Per-therapeutic PET/CT scans were delineated in consensus by two experienced physicians leading to the gold standard threshold to be applied. The mathematical expression of Thopt, the optimal threshold to be applied as a function of the maximum standard uptake value (SUVmax), was determined. The performance of this method (perPET-RT) was assessed by computing the DICE similarity coefficient (DSC) and was compared with 8 fixed threshold values and 3 adaptive thresholding methods. RESULTS: Thopt verified the following expression: Thopt = A.ln(1/SUVmax) + B where A and B were 2 constants. A and B were independent from the generation of PET/CT, but depended on the type of lesions (primary lung tumours vs. lymph nodes). PerPET-RT showed good to very good agreement in comparison to the gold standard. The mean and standard deviation of DSC value was 0.81 ± 0.13 for lung lesions and 0.78 ± 0.15 for lymph nodes. PerPET-RT showed a significant better agreement than the other segmentation methods (p < 0.001), except for one of the adaptive thresholding method ADT (p = 0.11). CONCLUSION: On the database used, perPET-RT has proven its reliability and accuracy for tumour delineation on per-therapeutic FDG PET/CT using only SUVmax measurement. This method may be used to delineate tumour volume for dose-escalation planning. TRIAL REGISTRATION: NCT01261598 , NCT01261585 , NCT01576796 .

Impact of consensus contours from multiple PET segmentation methods on the accuracy of functional volume delineation.

PURPOSE: The aim of this study was to evaluate the impact of consensus algorithms on segmentation results when applied to clinical PET images. In particular, whether the use of the majority vote or STAPLE algorithm could improve the accuracy and reproducibility of the segmentation provided by the combination of three semiautomatic segmentation algorithms was investigated. METHODS: Three published segmentation methods (contrast-oriented, possibility theory and adaptive thresholding) and two consensus algorithms (majority vote and STAPLE) were implemented in a single software platform (Artiview®). Four clinical datasets including different locations (thorax, breast, abdomen) or pathologies (primary NSCLC tumours, metastasis, lymphoma) were used to evaluate accuracy and reproducibility of the consensus approach in comparison with pathology as the ground truth or CT as a ground truth surrogate. RESULTS: Variability in the performance of the individual segmentation algorithms for lesions of different tumour entities reflected the variability in PET images in terms of resolution, contrast and noise. Independent of location and pathology of the lesion, however, the consensus method resulted in improved accuracy in volume segmentation compared with the worst-performing individual method in the majority of cases and was close to the best-performing method in many cases. In addition, the implementation revealed high reproducibility in the segmentation results with small changes in the respective starting conditions. There were no significant differences in the results with the STAPLE algorithm and the majority vote algorithm. CONCLUSION: This study showed that combining different PET segmentation methods by the use of a consensus algorithm offers robustness against the variable performance of individual segmentation methods and this approach would therefore be useful in radiation oncology. It might also be relevant for other scenarios such as the merging of expert recommendations in clinical routine and trials or the multiobserver generation of contours for standardization of automatic contouring.

Segmentation of heterogeneous or small FDG PET positive tissue based on a 3D-locally adaptive random walk algorithm.

A segmentation algorithm based on the random walk (RW) method, called 3D-LARW, has been developed to delineate small tumors or tumors with a heterogeneous distribution of FDG on PET images. Based on the original algorithm of RW [1], we propose an improved approach using new parameters depending on the Euclidean distance between two adjacent voxels instead of a fixed one and integrating probability densities of labels into the system of linear equations used in the RW. These improvements were evaluated and compared with the original RW method, a thresholding with a fixed value (40% of the maximum in the lesion), an adaptive thresholding algorithm on uniform spheres filled with FDG and FLAB method, on simulated heterogeneous spheres and on clinical data (14 patients). On these three different data, 3D-LARW has shown better segmentation results than the original RW algorithm and the three other methods. As expected, these improvements are more pronounced for the segmentation of small or tumors having heterogeneous FDG uptake.

[Interest of FDG-PET for lung cancer radiotherapy]. / Intérêt de la TEP au FDG pour la radiothérapie des cancers bronchiques.

The recent advances in medical imaging have profoundly altered the radiotherapy of non-small cell lung cancers (NSCLC). A meta-analysis has confirmed the superiority of FDG PET-CT over CT for initial staging. FDG PET-CT improves the reproducibility of target volume delineation, especially close to the mediastinum or in the presence of atelectasia. Although not formally validated by a randomized trial, the reduction of the mediastinal target volume, by restricting the irradiation to FDG-avid nodes, is widely accepted. The optimal method of delineation still remains to be defined. The role of FDG PET-CT in monitoring tumor response during radiotherapy is under investigation, potentially opening the way to adapting the treatment modalities to tumor radiation sensitivity. Other tracers, such as F-miso (hypoxia), are also under clinical investigation. To avoid excessive delays, the integration of PET-CT in routine practice requires quick access to the imaging equipment, technical support (fusion and image processing) and multidisciplinary delineation of target volumes.

Influence of PET/CT on radiologists and contrast-enhanced CT on nuclear medicine physicians in patients with lymphoma.

AIM: We assessed in this study the influence of contrast-enhanced CT (ceCT) on PET/CT interpretation and PET/CT on ceCT interpretation in patients with lymphoma, before and after chemotherapy. METHODS: Fifty patients with Hodgkin disease (N.=17) or non-Hodgkin lymphomas (N.=33) were assessed before and after chemotherapy. PET/CT were performed 60 minutes after injection of FDG. Iopamidol was then injected and followed, 50 seconds later, by another CT. PET images were successively reconstructed using non-enhanced CT (PET-) and ceCT (PET+). Four nuclear physicians rated PET- and PET+ in random order. Three radiologists initially rated ceCT alone and then ceCT along with PET+. RESULTS: Before chemotherapy, global agreement (GA) was 99% (k=0.96) when PET- was compared to PET+. Nine (5%) lesions were discordant, 5 according to PET- and 4 to PET+. After chemotherapy, GA was 99% (k=0.91). Eight (15%) lesions were discordant, 3 according to PET- and 5 to PET+. Before chemotherapy, GA was 97% (k=0.91) when ceCT was compared to ceCT with PET+. Twenty-one (12%) lesions were discordant, 16 when ceCT were analyzed alone and 5 when ceCT was analyzed with PET+. After chemotherapy, GA was 95% (k=0.76). All 30 (35%) discordant lesions were positive according to ceCT alone. A significant difference between the 2 procedures was found in the pelvis and in the groin (P<0.05). CONCLUSION: PET+ did not differ from PET-, before and after chemotherapy. Fewer abnormalities were observed, when ceCT was analyzed with PET+, particularly after chemotherapy, due to residual masses that are better analyzed with functional imaging.

Calcitriol (1,25-dihydroxycholecalciferol) enhances paclitaxel antitumor activity in vitro and in vivo and accelerates paclitaxel-induced apoptosis.

We demonstrated that calcitriol has antiproliferative activity in squamous cell carcinoma and prostatic adenocarcinoma and enhances the antitumor activity of platinum-based agents. In this study, we examined whether calcitriol also increases paclitaxel cytotoxicity. The effect of treatment on growth of the murine squamous cell carcinoma (SCCVII/SF) and human prostatic adenocarcinoma (PC-3) was determined by clonogenic assay, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and monitoring tumor growth. Treatment of SCC or PC-3 cells in vitro with calcitriol prior to paclitaxel significantly reduced clonogenic survival compared with either agent alone. Median-dose effect analysis revealed that calcitriol and paclitaxel interact synergistically. Treatment of SCC or PC-3 tumor-bearing mice with calcitriol prior to paclitaxel resulted in substantially greater growth inhibition than was achieved with either agent alone, supporting the combined use of calcitriol and paclitaxel in the treatment of solid tumors. To explore the molecular basis for the enhanced antitumor activity of this combination, the effect of treatment on p21(Waf-1) (p21), Bcl-2, and poly(ADP-ribose) polymerase expression was evaluated in PC-3. A 72-h pretreatment with calcitriol reduced p21 expression and increased paclitaxel cytotoxicity (measured after 24 h) without evidence of apoptosis [poly(ADP-ribose) polymerase cleavage]. After 48 h, paclitaxel induced apoptosis, the extent of which was increased similarly by pretreatment or concurrent treatment with calcitriol. We therefore propose a model for calcitriol enhancement of paclitaxel cytotoxicity in which the "early" (24 h) effects are schedule dependent and not attributed to enhancement of paclitaxel-induced apoptosis. In contrast, the "delayed" (48-h) enhancement of paclitaxel activity by calcitriol is schedule independent and associated with acceleration of apoptosis.

A novel approach for the identification of unique tumor vasculature binding peptides using an E. coli peptide display library.

BACKGROUND: Tumor neovascularization is necessary for continued tumor growth and metastasis. During the process of endothelial cell (EC) recruitment and tumor infiltration, specific molecular markers unique for this interaction are expressed on the EC surface. Targeting these molecular markers would, in effect, allow for specific tumor targeting. Tripeptide sequence motifs have previously been reported that will bind to angiogenic tumor ECs. These sequences were identified from in vivo phage peptide display libraries. The purpose of this study was to use a more simplified bacterial peptide display library in an in vitro system to seek out peptide motifs with unique binding to tumor microvasculature. METHODS: FliTrx is a bacterial peptide display library containing the entire repertoire of possible random dodecapeptides expressed on the flagella tip of E. coli. Two EC populations were used for the screening process, Matrigel invading cells (MAGIC) and tumor-derived endothelial cells (TDEC). MAGIC are obtained from ECs that infiltrate a subcutaneous fibroblast growth factor-containing Matrigel deposit, and TDEC are ECs selectively obtained from tumor vasculature. FliTrx cells were incubated with MAGIC at 4 degrees C to remove any potential clones displaying peptides that will bind to nonspecific EC surface targets. The non-binding cells were then incubated with TDEC, allowing for clones displaying potential binding peptides to bind tumor specific targets on TDECs. The bacterial population was then expanded and this "panning" process was carried out a total of five times. Peptide insert sequences from 100 bacterial colonies were analyzed for potential repetitive peptide motifs. RESULTS: Recurring peptide sequences were detected that were 3-mers (13 sequences) and 4-mers (4 sequences). Of the 3-mers, four repeated 3 times, whereas none of the 4-mers repeated more than twice. All of the repeated sequences were basic in charge, and arginine was the most commonly seen amino acid. A tripeptide basic-basic-nonpolar amino acid arrangement was the most prevalent charge sequence in all repetitive motifs (17 repeat sequences). Two test peptides showed TDEC binding specificity, and both conformed to the basic-basic-nonpolar motif. CONCLUSIONS: We report peptide sequences derived from panning an in vitro system designed to detect tumor-EC specific markers. These putative motifs may serve as molecular determinants for a novel therapeutic modality aimed at specifically targeting tumors through tumor angiogenic vessels.

1,25-Dihydroxycholecalciferol (1,25-D3) inhibits the growth of squamous cell carcinoma and down-modulates p21(Waf1/Cip1) in vitro and in vivo.

1,25-Dihydroxycholecalciferol (1,25-D3) has significant antitumor effects in the murine squamous cell carcinoma (SCC) tumor model in vitro and in vivo. We investigated the basis for this antiproliferative activity and found that, in vitro, 1,25-D3 administration is associated with altered expression of cell cycle regulatory proteins, treatment results in retinoblastoma dephosphorylation, decreased expression of p21(Waf1/Cip1) (p21) mRNA and protein, and increased expression of p27Kip1 (p27) mRNA and protein. Dexamethasone, which acts synergistically with 1,25-D3 to inhibit SCC proliferation, enhanced 1,25-D3-induced down-modulation of p21 without affecting the ability of 1,25-D3 to increase p27 expression. 1,25-D3 did not induce cleavage of poly(ADP-ribose) polymerase. These in vitro data suggest that 1,25-D3 exerts antitumor activity in SCC by perturbing cell cycle progression rather than by inducing apoptosis. In vivo, a 1,25-D3 treatment regimen that results in a decrease in SCC tumor volume is associated with a statistically significant decrease in intratumoral p21 expression. p21 expression is not changed in tumors isolated from control animals or animals treated with a nontherapeutic dose of 1,25-D3. Intratumoral p27 levels were not modulated by 1,25-D3 treatment. Thus, both in vitro and in vivo, 1,25-D3-mediated growth inhibition is associated with p21 down-modulation.

Enhancement of 1,25-dihydroxyvitamin D3-mediated antitumor activity with dexamethasone.

BACKGROUND: The active metabolite of vitamin D, i.e., 1,25-dihydroxycholecalciferol (1,25-D3), inhibits the growth of murine SCCVII/SF squamous cell carcinoma cells, both in vitro and in vivo. However, in vivo use of 1,25-D3 is hampered as a result of hypercalcemia (i.e., elevated levels of calcium in the blood). Glucocorticoids, such as dexamethasone, affect calcium absorption and modulate vitamin D receptor binding and have been used to treat hypercalcemia. In this study, we examined the effect of dexamethasone on tumor growth inhibition by 1,25-D3. METHODS: The effects of 1,25-D3 and dexamethasone, alone and in combination, on the growth of SCCVII/SF cells in in vitro culture or in vivo in female C3H/HeJ mice were determined by clonogenic tumor cell assay and/or by actual changes in tumor volume. Vitamin D receptor-ligand-binding activities in whole-cell extracts from cells (in culture), tumors, and normal tissues were assayed by single-point saturation analysis and equilibrium binding. RESULTS: Treatment of cultured SCCVII/SF cells with 500 nM dexamethasone for 24 hours before addition of 1,25-D3 reduced their survival. The growth of SCCVII/SF tumors was inhibited in mice treated simultaneously with dexamethasone and 1,25-D3 (as compared with no treatment or single-agent treatment); hypercalcemia was also reduced. Total vitamin D receptor content in SCCVII/SF cells was increased after treatment with dexamethasone. Treatment of tumor-bearing animals with dexamethasone (9 microg/day) for 7 days led to increased vitamin D receptor-ligand-binding activities in whole-cell extracts from tumor or kidneys and decreased activity in intestinal mucosa. CONCLUSIONS: Dexamethasone may enhance the antitumor effect of 1,25-D3 by increasing vitamin D receptor-ligand-binding activity.

Integrin activation suppresses etoposide-induced DNA strand breakage in cultured murine tumor-derived endothelial cells.

Tumor endothelium is critical for solid tumor growth and is a potential site for anticancer drug action. Within 2 h, etoposide caused marked DNA strand breakage in xenograft tumor-derived endothelial cells (TDECs). Etoposide-induced DNA breakage was inhibited by culturing TDECs on gelatin, type IV collagen, laminin, fibronectin, and the integrin ligand hexapeptide, GRGDSP, but not the inactive peptide, GRADSP. It was also inhibited when TDECs were on surfaces coated with antibodies to alpha 5, beta 1, or beta 3 integrin subunits and by clustering integrins with soluble antibodies. After 8 h with etoposide, TDECs detached from the monolayer, and 50-kb DNA fragments were seen. Fibronectin inhibited both processes. Thus, integrins are survival factors for TDEC that inhibit the genotoxicity of etoposide and may influence the sensitivity of tumors to drugs.

Interleukin 1 alpha and gamma-interferon induction of nitric oxide production from murine tumor-derived endothelial cells.

The role of nitric oxide (NO) in vascular function, host tumoricidal activity, and antiinflammatory effects is well documented. A number of cytokines induce NO from a variety of cell types. We have demonstrated in murine models that interleukin 1 alpha (IL-1 alpha) induces acute hemorrhagic necrosis, microvascular injury, and enhanced clonogenic tumor cell kill. Effects on the vasculature are observed only in tumor and not in normal tissues. Using methods established previously in our laboratory, murine tumor-derived and normal endothelial cells were cultured with IL-1 alpha, IFN-gamma, or IL-1 alpha/IFN-gamma at various doses with NO production quantitated through the measurement of nitrite by the Griess reaction. In tumor-derived endothelial cells, we demonstrated that neither cytokine alone was capable of inducing nitrite but that the combination of IL-1 alpha/IFN-gamma induced dose-dependent nitrite, with peak levels observed after 4 days incubation. When tumor-derived, normal yolk sac, mouse brain, or mouse aortic endothelial cells were treated with IL-1 alpha (100 units/ml)/IFN-gamma (10 units/ml), tumor-derived endothelial cells produced significantly more nitrite when compared to the normal endothelial cells. Nitrite production from IL-1 alpha/IFN-gamma was sensitive to the nitric oxide synthase inhibitors, NG-methyl-L-arginine or NG-nitro-L-arginine in a dose-dependent manner. In addition, dexamethasone significantly inhibited nitrite production from IL-1 alpha/IFN-gamma-treated, tumor-derived endothelial cells. These studies suggest that the antitumor activity of IL-1 alpha may be mediated through the production of NO from tumor-derived endothelial cells.

Potentiation by interleukin 1 alpha of cisplatin and carboplatin antitumor activity: schedule-dependent and pharmacokinetic effects in the RIF-1 tumor model.

We have previously demonstrated that the cytokine interleukin 1 alpha (IL-1 alpha) significantly potentiates the antitumor activity of a variety of chemotherapeutic agents, including cisplatin (cDDP). In studies described here, we examined the potential of combining IL-1 alpha and the platinum analogue carboplatin (CBDCA) and compared the schedule-dependent and pharmacokinetic effects for IL-1 alpha combinations with cDDP and CBDCA. RIF-1 tumor-bearing mice (C3H/HeJ) received i.p. injections of varying doses of CBDCA, alone or concurrently with IL-1 alpha (48 or 480 micrograms/kg). Clonogenic cell kill and tumor regrowth delay were significantly increased when CBDCA was combined with IL-1 alpha, at both doses, compared to either CBDCA or IL-1 alpha alone (P < 0.001 and P < 0.01, respectively). Although pretreatment with IL-1 receptor antagonist blocked the acute tumor hemorrhagic response induced by IL-1 alpha alone, IL-1 receptor antagonist only partially blocked IL-1 alpha enhancement of CBDCA or cDDP-mediated tumor cell kill. The IL-1 alpha enhancement of CBDCA-mediated tumor cell kill was highly schedule dependent, with maximum antitumor activity observed when IL-1 alpha was administered 4-12 h before CBDCA. In contrast, administration of IL-1 alpha from 24 h before or as late as 6 h after cDDP resulted in the same antitumor activity as simultaneous administration of cDDP and IL-1 alpha. Tumor and normal tissue platinum content were significantly increased by IL-1 alpha in animals treated with CBDCA (P < 0.01) but not in those treated with cDDP. The observed differences between cDDP and CBDCA may be explained by their known differential rates of clearance and protein binding affinities and are compatible with an induced alteration in CBDCA pharmacokinetics.