Gene therapy and cell therapy for the management of radiation damages to healthy tissues: Rationale and early results

International audienceNowadays, ionizing radiations have numerous applications, especially in medicine for diagnosis and therapy. Pharmacological radioprotection aims at increasing detoxification of free radicals. Radiomitigation aims at improving survival and proliferation of damaged cells. Both strategies are essential research area, as non-contained radiation can lead to harmful effects. Some advances allowing the comprehension of normal tissue injury mechanisms, and the discovery of related predictive biomarkers, have led to developing several highly promising radioprotector or radiomitigator drugs. Next to these drugs, a growing interest does exist for biotherapy in this field, including gene therapy and cell therapy through mesenchymal stem cells. In this review article, we provide an overview of the management of radiation damages to healthy tissues via gene or cell therapy in the context of radiotherapy. The early management aims at preventing the occurrence of these damages before exposure or just after exposure. The late management offers promises in the reversion of constituted late damages following irradiation

Physiopathologie et modulation pharmacologique de l’entérite radique

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Un traitement chronique à l'apeline chez la souris obèse et résistante à l'insuline augmente l'oxydation des acides gras et la biogénèse mitochondriale dans le muscle

Communication orale ; http://www.lesjfn.f

Nrf2 et Keap1 : biomarqueurs prédictifs de radiorésistance des cancers de la tête et du cou ?

International audienceObjectif de l’étudeDe nos jours, le pronostic des formes évoluées des carcinomes épidermoïdes de la tête et du cou reste défavorable. La radiorésistance intrinsèque de la tumeur en est en grande partie responsable. Plusieurs données suggèrent que les dérégulations du couple Keap1/Nrf2, entraînant une augmentation de l’expression de certains gènes cytoprotecteurs, pourraient avoir un rôle clé dans les phénomènes de radiorésistance. L’objectif de cette étude était d’analyser la corrélation potentielle entre la radiosensibilité et l’expression du couple Nrf2/Keap1, proposant alors ces protéines comme des biomarqueurs prédictifs de radiosensibilité des carcinomes épidermoïdes de la tête et du cou.Matériel et méthodeNous avons sélectionné cinq lignées cellulaires humaines de carcinome épidermoïde de la tête et du cou présentant autant de degrés différents de radiorésistance. La radiorésistance est définie, conventionnellement, par la proportion de survie (test clonogénique de mort cellulaire) après une irradiation de 2 Gy (survival fraction 2 : SF2). Le niveau d’expression des protéines Keap1 et Nrf2 a été analysé par westernblot.RésultatUne nette corrélation a été établie entre le degré de radiorésistance des différentes lignées et l’expression de la protéine Keap1 (R2 = 0,94). En revanche, il n’y avait pas de corrélation avec la protéine Nrf2 (R2 = 0,02).ConclusionCette étude préliminaire, réalisée in vitro, propose Keap1 comme un nouveau bio-marqueur de radiorésistance et offre ainsi une nouvelle perspective dans le choix du plan de traitement des carcinomes épidermoïdes de la tête et du cou. En effet, connaissant les degrés de radiorésistance de leur tumeur, les oncologues pourraient engager les patients, dès le diagnostic, vers d’autres stratégies thérapeutiques. Ces résultats préliminaires doivent maintenant être confirmés par l’examen de niveau Keap1 dans les biopsies de carcinomes épidermoïdes de la tête et du cou afin d’évaluer leur éventuelle implication future dans la décision clinique du plan de traitement

Ciblage de la famille HER dans les cancers ORL : efficacité biologique de l’association de cétuximab et de pertuzumab combinée à l’irradiation photonique

International audienceObjectif de l’étudeLes cancers épidermoïdes ORL sont associés à un fort taux de récidive loco-régionale et métastatique. Les cellules souches cancéreuses, sous-population hautement migratoire, semblent être une hypothèse majeure à l’origine de la résistance aux traitements. L’objectif du travail était de comparer l’efficacité du blocage pan-HER par une association cétuximab-pertuzumab associé à l’irradiation photonique dans les processus d’invasion et migration de la lignée SQ20B et sa sous-population souche.Matériel et méthodeLa sous-population de CSCs de la lignée SQ20B a été isolée par double tri selon les critères SQ20B/SP/CD44High. La prolifération des cellules SQ20B et SQ20B/CSCs a été étudiée après traitement par cétuximab 5 nM et/ou pertuzumab 20 μg/mL avec ou sans irradiation photonique à 10 Gy. L’analyse de la migration et de l’invasion a été réalisée par test de blessure avec et sans matrigel (IncuCyte). L’activation de récepteur de l’epidermal growth factor (EGFR) (Tyr1068) et des voies de signalisation intracellulaires (phospho-AKT et phospho-MEK1/2) a été étudiée en réponse aux traitements par western blot (WES).RésultatsLe cétuximab inhibe la prolifération cellulaire des cellules SQ20B et non celle de la sous-population souche. L’association cétuximab-pertuzumab inhibe significativement la prolifération des cellules SQ20B et SQ20B/CSCs. La double association cétuximab-pertuzumab associée à une irradiation de 10 Gy inhibe significativement la migration et l’invasion des deux populations cellulaires. Le double traitement inhibe la phosphorylation d’EGFR dans les deux populations. Les cellules SQ20B expriment fortement phospho-AKT à l’inverse des SQ20B/CSCs qui expriment phospho-MEK1/2. Enfin, l’association cétuximab-pertuzumab-10 Gy inhibe fortement l’expression de phospho-AKT et phospho-MEK1/2.ConclusionLe double traitement par cétuximab-pertuzumab associé à l’irradiation photonique inhibe significativement la prolifération, la migration et l’invasion de la lignée SQ20B et sa sous-population souche

Effects of Dietary Eicosapentaenoic Acid (EPA) Supplementation in High-Fat Fed Mice on Lipid Metabolism and Apelin/APJ System in Skeletal Muscle

Various studies have shown that eicosapentaenoic acid (EPA) has beneficial effects on obesity and associated disorders. Apelin, the ligand of APJ receptor also exerts insulin-sensitizing effects especially by improving muscle metabolism. EPA has been shown to increase apelin production in adipose tissue but its effects in muscle have not been addressed. Thus, the effects of EPA supplementation (36 g/kg EPA) in high-fat diet (HFD) (45% fat, 20% protein, 35% carbohydrate) were studied in mice with focus on muscle lipid metabolism and apelin/APJ expression. Compared with HFD mice, HFD+EPA mice had significantly less weight gain, fat mass, lower blood glucose, insulinemia and hepatic steatosis after 10 weeks of diet. In addition, EPA prevented muscle metabolism alterations since intramuscular triglycerides were decreased and b-oxidation increased. In soleus muscles of HFD+EPA mice, apelin and APJ expression were significantly increased compared to HFD mice. However, plasma apelin concentrations in HFD and HFD+EPA mice were similar. EPA-induced apelin expression was confirmed in differentiated C2C12 myocytes but in this model, apelin secretion was also increased in response to EPA treatment. In conclusion, EPA supplementation in HFD prevents obesity and metabolic alterations in mice, especially in skeletal muscle. Since EPA increases apelin/APJ expression in muscle, apelin may act in a paracrine/autocrine manner to contribute to these benefical effect

HER Family Blockade in Head and Neck Squamous Cell Carcinoma: Couple Therapy Efficacy of Cetuximab and Pertuzumab Combined With Photon Irradiation

International audiencePurpose/Objective(s)Head and neck squamous cell carcinoma (HNSCC) is a malignancy still associated with severe mortality, due to loco-regional recurrences or distant metastasis. Head and neck cancer stem cells (CSCs) are highly resistant to treatment and have large migratory abilities. These particular properties could explain treatment resistances in this location. If EGFR is strongly overexpressed in 80-100% of HNSCC, HER2 and HER3 seem to be also expressed in these lines. The aim of the present study was to explore the efficacy of HER1-2-3 blockade through cetuximab-pertuzumab association with or without photon irradiation on the proliferation and migration/invasion capabilities of a HNSCC chemo and radio resistant human cell line (SQ20B) and its corresponding stem cell line (SQ20B/CSCs).Materials/MethodsSQ20B/CSCs subpopulation was isolated through double cell sorting: side population (SP) (Hoechst exclusion) and CD44 staining: SP/CD44High. SQ20B and SQ20B/CSCs proliferation was studied after treatment with cetuximab 5nM + pertuzumab 20mg/mL treatment +/- 10Gy photon irradiation. Invasion and migration were assessed with scratch wound assay with or without matrigel. EGFR, phospho-EGFR (Tyr1068), HER2 and HER3 basal protein expression was studied. Activation or inhibition of RAS/MAPK and AKT-mTOR downstream signaling cascade was studied through phospho-AKT (Ser473), phospho-MEK1/2(Ser217/221) expression exposed to combined treatments.ResultsCetuximab strongly inhibits SQ20B proliferation, migration and invasion when it has a small effect on SQ20B/CSCs. Cetuximab-pertuzumab treatment combined with radiation has a potent significant inhibitory effect on SQ20B and SQ20B/CSCs proliferation, migration and invasion. EGFR is overexpressed in SQ20B, and under-expressed in SQ20B/CSCs, while HER2 and HER3 are expressed equivalently in both populations in basal conditions. Phospho-AKT is strongly expressed in SQ20B, at the opposite of SQ20B/CSCs which express phospho-MEK1/2. Cetuximab-pertuzumab treatment combination with 10Gy photon irradiation switches off both phospho-AKT and phospho-MEK1/2 expression in the two populations.ConclusionCetuximab-pertuzumab couple pan-HER treatment combined with photon irradiation significantly inhibits proliferation, invasion and migration of SQ20B HNSCC cell line and its CSCs subpopulation, through both AKT-mTOR and Ras-MAPK downstream signaling blockade. HER family seems to be a promising therapeutic target in HNSCC

Biomarkers of resistance to radiation therapy: a prospective study in cervical carcinoma

Abstract Background Clinical parameters and proteins have recently been suggested as possible causes of radiotherapy (RT) resistance in cervical carcinoma (CC). The objective of the present study was to validate prognostic biomarkers of radiation resistance. Methods The present prospective study included patients undergoing RT with curative intent for histologically proven locally advanced squamous cell CC. Tissues and blood samples were systematically collected before RT initiation. Immuno-histochemistry was performed (IGF-IR α and β, GAPDH, HIF-1 alpha, Survivin, GLUT1, CAIX, hTERT and HKII). Response to radiation was assessed through tumour response 3 months after RT completion, through overall survival (OS) and through progression-free survival (PFS). Results One hundred forty nine patients with a mean age of 46 years were included, with FIGO IIB (n = 53) and FIGO IIIB (n = 96) CCs. 61 patients were treated with exclusive RT + brachytherapy and 88 underwent chemo-radiotherapy + brachytherapy. Our findings suggest an association between hemoglobin level (Hb) (>11 g/dL) and 3 months complete response (p = 0.02). Hb level < 11 g/dL was associated with decreased PFS (p = 0.05) and OS (p = 0.08). Overexpression of IGF-1R β was correlated with a decreased OS (p = 0.007). Overexpression of GLUT1 was marginally correlated with reduced OS (p = 0.05). PFS and OS were significantly improved in patients undergoing chemoradiation versus exclusive radiotherapy (PFS: p = 0.04; OS: p = 0.01). Conclusions IGF-1R β overexpression and Hb level (≤11 g/dl) were associated with poor prognosis, and thus appear to be possible interesting biomarkers of radiation resistance. Our results corroborate previous pre-clinical studies suggesting IGF-1R and hypoxia to be part of the biological pathways leading to radio-resistance

Targeting Cancer Stem Cells in HNSCC: Synergic Effect of Cetuximab and ABT-199 in Combination with Photon Radiation

International audiencePurpose/Objective(s)Concurrent cetuximab based radio-chemotherapy is a validated scheme in head and neck squamous cell carcinoma (HNSCC). Cancer Stem Cells (CSCs) are highly resistant to treatment, have large migratory abilities, and are hypothesized to be responsible for a significant part of recurrences. Apoptotic signaling in CSCs is a major way of treatment escape, through Bcl-2 proteins family. The aim of the present study was to explore the synergic effect between cetuximab and an anti-Bcl-2 antibody (ABT-199), when combined or not with photon radiation.Materials/MethodsHNSCC chemo and radio resistant human cell line (SQ20B) and its corresponding stem cell line (SQ20B/CSCs) were used to test the treatment combinations. HaCaT cell line was used to assess toxicity on healthy cell population. SQ20B/CSCs subpopulation was isolated through double cell sorting: side population (SP) (Hoechst exclusion) and CD44 staining: SP/CD44High. SQ20B and SQ20B/CSCs proliferation, invasion/migration, and apoptosis were studied after an exposition to cetuximab 5nM + ABT-199 10mM treatment +/- 10Gy photon irradiation. Invasion and migration were assessed based on scratch wound assay with or without matrigel. Apoptosis was measured using caspases 3/7. 3D spheroid assay was performed to validate the results in a 3D culture approach. EGFR, phospho-EGFR (Tyr1068), Bcl-2 and Bcl-xl protein expression were studied.ResultsCetuximab strongly inhibited SQ20B proliferation, migration and invasion whereas it had little effect on SQ20B/CSCs. Conversely, ABT-199 significantly inhibited these properties on SQ20B/CSCs, without showing any effect on SQ20B parental cell line. Cetuximab-ABT-199 combined with radiation had a significant inhibitory effect on both SQ20B and SQ20B/CSCs proliferation, migration and invasion. Although exclusive cetuximab had no pro-apoptotic effect, activation of caspases 3/7 was induced by ABT-199 and enhanced by the cetuximab+ABT-199 combination in both populations. Cetuximab was a strong inhibitor of 3D-spheroid proliferation in SQ20B, whereas ABT-199 strongly decreased spheroid size in CSCs. EGFR was overexpressed in SQ20B, and under-expressed in SQ20B/CSCs. Bcl2 was overexpressed in SQ20B/CSCs. Although cetuximab moderately inhibited HaCaT cell proliferation, the drug combination did not significantly enhanced toxicity.ConclusionCetuximab+ABT-199 combined with photon radiation significantly inhibited proliferation, invasion and migration of SQ20B HNSCC cell line and its CSCs subpopulation, with an acceptable toxicity profile on healthy cell lines. Apoptotic cell death was enhanced by this drug combination

Navigating the highlights of phase III trials: a watchful eye on evidence-based radiotherapy

International audienceBackground: Phase III randomized controlled trials (RCTs) are the cornerstone of evidence-based oncology. However, there is no exhaustive review describing the radiotherapy RTCs characteristics. The objective of the present study was to describe features of all phase III RCTs including at least a radiation therapy. Methods and materials: Requests were performed in the Medline database (via PubMed). The latest update was performed in April 2016, using the following MESH terms: 'clinical trials: phase III as topic', 'radiotherapy', 'brachytherapy', as keywords. Results: A total of 454 phase III RCTs were identified. Studies were mainly based on open (92.1%) multicenter (77.5%) designs, analyzed in intend to treat (67.6%), aiming at proving superiority (91.6%) through overall survival assessment (46.5%). Most frequently studied malignancies were head and neck (21.8%), lung (14.3%) and prostate cancers (9.9%). Patients were mainly recruited with a locally advanced disease (73.7%). Median age was 59 years old. Out of 977 treatment arms, 889 arms experienced radiotherapy, mainly using 3D-conformal radiotherapy (288 arms, 32.4%). Intensity-modulated techniques were tested in 12 arms (1.3%). The intervention was a non-cytotoxic agent addition in 89 studies (19.6%), a radiation dose/ fractionation modification in 74 studies (16.3%), a modification of chemotherapy regimen in 63 studies (13.9%), a chemotherapy addition in 63 studies (13.9%) and a radiotherapy addition in 53 trials (11.7%). With a median follow-up of 50 months, acute all-grade and grade 3–5 toxicities were reported in 49.6% and 69.4% of studies, respectively. Radiotherapy technique, follow-up and late toxicities were reported in 60.1%, 74%, and 31.1% of studies, respectively. Conclusion: Phase III randomized controlled trials featured severe limitations, since a third did not report radiotherapy technique, follow-up or late toxicities. The fast-paced technological evolution creates a discrepancy between literature and radiotherapy techniques performed in daily-routine, suggesting that phase III methodology needs to be reinvented