RESUMO
PURPOSE: Tumor hypoxia fuels an aggressive tumor phenotype and confers resistance to anticancer treatments. We conducted a clinical trial to determine whether the antimalarial drug atovaquone, a known mitochondrial inhibitor, reduces hypoxia in non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: Patients with NSCLC scheduled for surgery were recruited sequentially into two cohorts: cohort 1 received oral atovaquone at the standard clinical dose of 750 mg twice daily, while cohort 2 did not. Primary imaging endpoint was change in tumor hypoxic volume (HV) measured by hypoxia PET-CT. Intercohort comparison of hypoxia gene expression signatures using RNA sequencing from resected tumors was performed. RESULTS: Thirty patients were evaluable for hypoxia PET-CT analysis, 15 per cohort. Median treatment duration was 12 days. Eleven (73.3%) atovaquone-treated patients had meaningful HV reduction, with median change -28% [95% confidence interval (CI), -58.2 to -4.4]. In contrast, median change in untreated patients was +15.5% (95% CI, -6.5 to 35.5). Linear regression estimated the expected mean HV was 55% (95% CI, 24%-74%) lower in cohort 1 compared with cohort 2 (P = 0.004), adjusting for cohort, tumor volume, and baseline HV. A key pharmacodynamics endpoint was reduction in hypoxia-regulated genes, which were significantly downregulated in atovaquone-treated tumors. Data from multiple additional measures of tumor hypoxia and perfusion are presented. No atovaquone-related adverse events were reported. CONCLUSIONS: This is the first clinical evidence that targeting tumor mitochondrial metabolism can reduce hypoxia and produce relevant antitumor effects at the mRNA level. Repurposing atovaquone for this purpose may improve treatment outcomes for NSCLC.
Assuntos
Atovaquona/farmacologia , Regulação Neoplásica da Expressão Gênica , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Hipóxia Tumoral/efeitos dos fármacos , Hipóxia Tumoral/genética , Atovaquona/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Metabolismo Energético , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Feminino , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Neoplasias Pulmonares/diagnóstico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Imagem Molecular , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Fator de Transcrição STAT3/metabolismoRESUMO
BACKGROUND: Pre-clinically, phosphoinositide 3-kinase (PI3K) inhibition radiosensitises tumours by increasing intrinsic radiosensitivity and by reducing tumour hypoxia. We assessed whether buparlisib, a class 1 PI3K inhibitor, can be safely combined with radiotherapy in patients with non-small cell lung carcinoma (NSCLC) and investigated its effect on tumour hypoxia. METHODS: This was a 3 + 3 dose escalation and dose expansion phase I trial in patients with advanced NSCLC. Buparlisib dose levels were 50 mg, 80 mg and 100 mg once daily orally for 2 weeks, with palliative thoracic radiotherapy (20 Gy in 5 fractions) delivered during week 2. Tumour hypoxic volume (HV) was measured using 18F-fluoromisonidazole positron-emission tomography-computed tomography at baseline and following 1 week of buparlisib. RESULTS: Twenty-one patients were recruited with 9 patients evaluable for maximum tolerated dose (MTD) analysis. No dose-limiting toxicity was reported; therefore, 100 mg was declared the MTD, and 10 patients received this dose in the expansion phase. Ninety-four percent of treatment-related adverse events were ≤grade 2 with fatigue (67%), nausea (24%) and decreased appetite (19%) most common per patient. One serious adverse event (grade 3 hypoalbuminaemia) was possibly related to buparlisib. No unexpected radiotherapy toxicity was reported. Ten (67%) of 15 patients evaluable for imaging analysis were responders with 20% median reduction in HV at the MTD. CONCLUSION: This is the first clinical trial to combine a PI3K inhibitor with radiotherapy in NSCLC and investigate the effects of PI3K inhibition on tumour hypoxia. This combination was well tolerated and PI3K inhibition reduced hypoxia, warranting investigation into whether this novel class of radiosensitisers can improve radiotherapy outcomes.
Assuntos
Adenocarcinoma de Pulmão/terapia , Aminopiridinas/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/terapia , Carcinoma de Células Escamosas/terapia , Neoplasias Pulmonares/terapia , Morfolinas/uso terapêutico , Inibidores de Fosfoinositídeo-3 Quinase/uso terapêutico , Radiossensibilizantes/uso terapêutico , Hipóxia Tumoral , Adenocarcinoma de Pulmão/diagnóstico por imagem , Adenocarcinoma de Pulmão/metabolismo , Idoso , Anorexia/induzido quimicamente , Carcinoma Pulmonar de Células não Pequenas/diagnóstico por imagem , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma de Células Escamosas/diagnóstico por imagem , Carcinoma de Células Escamosas/metabolismo , Quimiorradioterapia , Fadiga/induzido quimicamente , Feminino , Humanos , Neoplasias Pulmonares/diagnóstico por imagem , Neoplasias Pulmonares/metabolismo , Masculino , Dose Máxima Tolerável , Pessoa de Meia-Idade , Misonidazol/análogos & derivados , Náusea/induzido quimicamente , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , RadioterapiaRESUMO
Cyclin-dependent kinase 1 (CDK1) orchestrates the transition from the G2 phase into mitosis and as cancer cells often display enhanced CDK1 activity, it has been proposed as a tumor specific anti-cancer target. Here we show that the effects of CDK1 inhibition are not restricted to tumor cells but can also reduce viability in non-cancer cells and sensitize them to radiation in a cell cycle dependent manner. Radiosensitization by the specific CDK1 inhibitor, RO-3306, was determined by colony formation assays in three tumor lines (HeLa, T24, SQ20B) and three non-cancer lines (HFL1, MRC-5, RPE). Initial results showed that CDK1 inhibition radiosensitized tumor cells, but did not sensitize normal fibroblasts and epithelial cells in colony formation assays despite effective inhibition of CDK1 signaling. Further investigation showed that normal cells were less sensitive to CDK1 inhibition because they remained predominantly in G1 for a prolonged period when plated in colony formation assays. In contrast, inhibiting CDK1 a day after plating, when the cells were going through G2/M phase, reduced their clonogenic survival both with and without radiation. Our finding that inhibition of CDK1 can damage normal cells in a cell cycle dependent manner indicates that targeting CDK1 in cancer patients may lead to toxicity in normal proliferating cells. Furthermore, our finding that cell cycle progression becomes easily stalled in non-cancer cells under normal culture conditions has general implications for testing anti-cancer agents in these cells.
Assuntos
Proteína Quinase CDC2/antagonistas & inibidores , Divisão Celular/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Fase G2/efeitos dos fármacos , Radiossensibilizantes/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Células HeLa , Humanos , Transdução de Sinais/efeitos dos fármacosRESUMO
The nuclear pore complex (NPC) machinery is emerging as an important determinant in the maintenance of genome integrity and sensitivity to DNA double-strand break (DSB)-inducing agents, such as ionising radiation (IR). In this study, using a high-throughput siRNA screen, we identified the central channel NPC protein Nup54, and concomitantly its molecular partners Nup62 and Nup58, as novel factors implicated in radiosensitivity. Nup54 depletion caused an increase in cell death by mitotic catastrophe after IR, and specifically enhanced both the duration of the G2 arrest and the radiosensitivity of cells that contained replicated DNA at the time of IR exposure. Nup54-depleted cells also exhibited increased formation of chromosome aberrations arisen from replicated DNA. Interestingly, we found that Nup54 is epistatic with the homologous recombination (HR) factor Rad51. Moreover, using specific DNA damage repair reporters, we observed a decreased HR repair activity upon Nup54 knockdown. In agreement with a role in HR repair, we also demonstrated a decreased formation of HR-linked DNA synthesis foci and sister chromatid exchanges after IR in cells depleted of Nup54. Our study reveals a novel role for Nup54 in the response to IR and the maintenance of HR-mediated genome integrity.
Assuntos
Replicação do DNA , DNA/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/metabolismo , Reparo de DNA por Recombinação , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Sobrevivência Celular/efeitos da radiação , DNA/genética , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Células HeLa , Humanos , Células MCF-7 , Poro Nuclear/genética , Poro Nuclear/metabolismo , Complexo de Proteínas Formadoras de Poros Nucleares/genética , Interferência de RNA , Rad51 Recombinase/genética , Rad51 Recombinase/metabolismo , Radiação Ionizante , Troca de Cromátide Irmã/efeitos da radiaçãoRESUMO
PURPOSE: To investigate the induction of type III interferons (IFNs) in human cancer cells by gamma-rays. METHODS AND MATERIALS: Type III IFN expression in human cancer cell lines after gamma-ray irradiation in vitro was assessed by reverse transcription-quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. Signaling pathways mediating type III IFN induction were examined by a variety of means, including immunoblotting, flow cytometry, confocal imaging, and reverse transcription-quantitative polymerase chain reaction. Key mediators in these pathways were further explored and validated using gene CRISPR knockout or short hairpin RNA knockdown. RESULTS: Exposure to gamma-rays directly induced type III IFNs (mainly IFNL1) in human cancer cell lines in dose- and time-dependent fashions. The induction of IFNL1 was primarily mediated by the cytosolic DNA sensors-STING-TBK1-IRF1 signaling axis, with a lesser contribution from the nuclear factor kappa b signaling in HT29 cells. In addition, type III IFN signaling through its receptors serves as a positive feedback loop, further enhancing IFN expression via up-regulation of the kinases in the STING-TBK1 signaling axis. CONCLUSIONS: Our results suggest that IFNL1 can be up-regulated in human cancer cell lines after gamma-ray treatment. In HT29 cells this induction occurs via the STING pathway, adding another layer of complexity to the understanding of radiation-induced antitumor immunity, and may provide novel insights into IFN-based cancer treatment.
Assuntos
Raios gama/uso terapêutico , Fator Regulador 1 de Interferon/metabolismo , Interleucinas/efeitos da radiação , Proteínas de Membrana/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Relação Dose-Resposta à Radiação , Edição de Genes/métodos , Técnicas de Inativação de Genes , Células HT29 , Células HeLa , Humanos , Interferons , Interleucinas/metabolismo , NF-kappa B/metabolismo , RNA Interferente Pequeno , Receptores de Interferon/metabolismo , Regulação para CimaRESUMO
Cancer cells have upregulated glycolysis compared with normal cells, which has led many to the assumption that oxidative phosphorylation (OXPHOS) is downregulated in all cancers. However, recent studies have shown that OXPHOS can be also upregulated in certain cancers, including leukemias, lymphomas, pancreatic ductal adenocarcinoma, high OXPHOS subtype melanoma, and endometrial carcinoma, and that this can occur even in the face of active glycolysis. OXPHOS inhibitors could therefore be used to target cancer subtypes in which OXPHOS is upregulated and to alleviate therapeutically adverse tumor hypoxia. Several drugs including metformin, atovaquone, and arsenic trioxide are used clinically for non-oncologic indications, but emerging data demonstrate their potential use as OXPHOS inhibitors. We highlight novel applications of OXPHOS inhibitors with a suitable therapeutic index to target cancer cell metabolism. Clin Cancer Res; 24(11); 2482-90. ©2018 AACR.
Assuntos
Terapia de Alvo Molecular , Neoplasias/metabolismo , Neoplasias/terapia , Fosforilação Oxidativa , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Biomarcadores , Metabolismo Energético/efeitos dos fármacos , Humanos , Redes e Vias Metabólicas , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Terapia de Alvo Molecular/métodos , Neoplasias/etiologia , Fosforilação Oxidativa/efeitos dos fármacosRESUMO
PURPOSE: The aim of this study was to determine the relative abilities of compartment models to describe time-courses of 18 F-fluoromisonidazole (FMISO) uptake in tumor voxels of patients with non-small cell lung cancer (NSCLC) imaged using dynamic positron emission tomography. Also to use fits of the best-performing model to investigate changes in fitted rate-constants with distance from the tumor edge. METHODS: Reversible and irreversible two- and three-tissue compartment models were fitted to 24 662 individual voxel time activity curves (TACs) obtained from tumors in nine patients, each imaged twice. Descriptions of the TACs provided by the models were compared using the Akaike and Bayesian information criteria (AIC and BIC). Two different models (two- and three-tissue) were fitted to 30 measured voxel TACs to provide ground-truth TACs for a statistical simulation study. Appropriately scaled noise was added to each of the resulting ground-truth TACs, generating 1000 simulated noisy TACs for each ground-truth TAC. The simulation study was carried out to provide estimates of the accuracy and precision with which parameter values are determined, the estimates being obtained for both assumptions about the ground-truth kinetics. A BIC clustering technique was used to group the fitted rate-constants, taking into consideration the underlying uncertainties on the fitted rate-constants. Voxels were also categorized according to their distance from the tumor edge. RESULTS: For uptake time-courses of individual voxels an irreversible two-tissue compartment model was found to be most precise. The simulation study indicated that this model had a one standard deviation precision of 39% for tumor fractional blood volumes and 37% for the FMISO binding rate-constant. Weighted means of fitted FMISO binding rate-constants of voxels in all tumors rose significantly with increasing distance from the tumor edge, whereas fitted fractional blood volumes fell significantly. When grouped using the BIC clustering, many centrally located voxels had high-fitted FMISO binding rate-constants and low rate-constants for tracer flow between the vasculature and tumor, both indicative of hypoxia. Nevertheless, many of these voxels had tumor-to-blood (TBR) values lower than the 1.4 level commonly expected for hypoxic tissues, possibly due to the low rate-constants for tracer flow between the vasculature and tumor cells in these voxels. CONCLUSIONS: Time-courses of FMISO uptake in NSCLC tumor voxels are best analyzed using an irreversible two-tissue compartment model, fits of which provide more precise parameter values than those of a three-tissue model. Changes in fitted model parameter values indicate that levels of hypoxia rise with increasing distance from tumor edges. The average FMISO binding rate-constant is higher for voxels in tumor centers than in the next tumor layer out, but the average value of the more simplistic TBR metric is lower in tumor centers. For both metrics, higher values might be considered indicative of hypoxia, and the mismatch in this case is likely to be due to poor perfusion at the tumor center. Kinetics analysis of dynamic PET images may therefore provide more accurate measures of the hypoxic status of such regions than the simpler TBR metric, a hypothesis we are presently exploring in a study of tumor imaging versus histopathology.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/diagnóstico por imagem , Neoplasias Pulmonares/diagnóstico por imagem , Tomografia por Emissão de Pósitrons , Teorema de Bayes , Humanos , Cinética , Misonidazol/análogos & derivados , Misonidazol/farmacocinética , Radiossensibilizantes/farmacocinética , Compostos RadiofarmacêuticosRESUMO
The identification of genetic determinants that underpin tumor radioresistance can help the development of targeted radiosensitizers or aid personalization of radiotherapy treatment. Here we identify signal recognition particle 72kDa (SRP72) as a novel gene involved in radioresistance. Knockdown of SRP72 resulted in significant radiosensitization of HeLa (cervical), PSN-1 (pancreatic), and T24 (bladder), BT-549 (breast) and MCF7 (breast) tumor lines as measured by colony formation assays. SRP72 depletion also resulted in the radiosensitization of normal lung fibroblast cell lines (HFL1 and MRC-5), demonstrating that the effect is not restricted to tumor cells. Increased radiosensitivity was not due to impaired DNA damage signaling or repair as assessed by γ-H2AX foci formation. Instead SRP72 depletion was associated with elevated levels of apoptosis after irradiation, as measured by caspase 3/7 activity, PARP-cleavage and Annexin-V staining, and with an induction of the unfolded protein response. Together, our results show that SRP72 is a novel gene involved in radioresistance.
Assuntos
Tolerância a Radiação , Partícula de Reconhecimento de Sinal/genética , Apoptose , Sobrevivência Celular/efeitos da radiação , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Células MCF-7 , Biossíntese de Proteínas , Partícula de Reconhecimento de Sinal/metabolismo , Resposta a Proteínas não DobradasRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is considered a non-immunogenic tumor, and immune checkpoint inhibitor monotherapy lacks efficacy in this disease. Radiotherapy (RT) can stimulate the immune system. Here, we show that treatment of KPC and Pan02 murine PDAC cells with RT and gemcitabine upregulated PD-L1 expression in a JAK/Stat1-dependent manner. In vitro, PD-L1 inhibition did not alter radio- and chemosensitivity. In vivo, addition of anti-PD-L1 to high (12, 5 × 3, 20 Gy) but not low (6, 5 × 2 Gy) RT doses significantly improved tumor response in KPC and Pan02 allografts. Radiosensitization after PD-L1 blockade was associated with reduced CD11b+Gr1+ myeloid cell infiltration and enhanced CD45+CD8+ T-cell infiltration with concomitant upregulation of T-cell activation markers including CD69, CD44, and FasL, and increased CD8:Treg ratio. Depletion of CD8+ T cells abrogated radiosensitization by anti-PD-L1. Blockade of PD-L1 further augmented the effect of high RT doses (12 Gy) in preventing development of liver metastases. Exploring multiple mathematical models reveals a mechanism able to explain the observed synergy between RT and anti-PD-L1 therapy. Our findings provide a rationale for testing the use of immune checkpoint inhibitors with RT in PDAC.
Assuntos
Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/radioterapia , Antígeno B7-H1/antagonistas & inibidores , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/radioterapia , Desoxicitidina/análogos & derivados , Radiossensibilizantes/administração & dosagem , Animais , Linfócitos T CD8-Positivos , Desoxicitidina/administração & dosagem , Modelos Animais de Doenças , Camundongos , Modelos Teóricos , Resultado do Tratamento , GencitabinaRESUMO
We assessed the prognostic value of hypoxia (carbonic anhydrase 9; CA9), vessel density (CD31), with macrophages (CD68) and B cells (CD20) that can interact and lead to immune suppression and disease progression using scanning and histological mapping of whole-mount FFPE pancreatectomy tissue sections from 141 primarily resectable pancreatic ductal adenocarcinoma (PDAC) samples treated with surgery and adjuvant chemotherapy. Their expression was correlated with clinicopathological characteristics, and overall survival (OS), progression-free survival (PFS), local progression-free survival (LPFS) and distant metastases free-survival (DMFS), also in the context of stroma density (haematoxylin-eosin) and activity (alpha-smooth muscle actin). The median OS was 21 months after a mean follow-up of 20 months (range, 2-69 months). The median tumor surface area positive for CA9 and CD31 was 7.8% and 8.1%, respectively. Although total expression of these markers lacked prognostic value in the entire cohort, nevertheless, high tumor compartment CD68 expression correlated with worse PFS (p = 0.033) and DMFS (p = 0.047). Also, high CD31 expression predicted for worse OS (p = 0.004), PFS (p = 0.008), LPFS (p = 0.014) and DMFS (p = 0.004) in patients with moderate density stroma. High stromal and peripheral compartment CD68 expression predicted for significantly worse outcome in patients with loose and moderate stroma density, respectively. Altogether, in contrast to the current notion, hypoxia levels in PDAC appear to be comparable to other malignancies. CD31 and CD68 constitute prognostic markers in patient subgroups that vary according to tumor compartment and stromal density. Our study provides important insight on the pathophysiology of PDAC and should be exploited for future treatments.
Assuntos
Antígenos CD20/metabolismo , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Antígenos de Neoplasias/metabolismo , Anidrase Carbônica IX/metabolismo , Carcinoma Ductal Pancreático/metabolismo , Neoplasias Pancreáticas/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Células Estromais/metabolismo , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/terapia , Terapia Combinada , Feminino , Humanos , Hipóxia/metabolismo , Imuno-Histoquímica , Macrófagos/metabolismo , Masculino , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Neovascularização Patológica/metabolismo , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/terapia , Prognóstico , Análise de Sobrevida , Neoplasias PancreáticasRESUMO
AIMS: We have previously shown in a phase I trial that nelfinavir (NFV) is safe with chemoradiation in PDAC with good signs for efficacy. Reverse translationally, we aimed to test the influence of PSCs on nelfinavir mediated radiosensitization to PDAC preclinically, because PDAC is very rich in desmoplasia and PSCs are known to mediate radioresistance. METHODS: In a direct co-culture model of several PDAC cell lines with PSC we performed clonogenic assays +/- nelfinavir. This was repeated exposing cells to hypoxic conditions. In xenograft PDAC tumors we tested radiation +/- nelfinavir +/- PSC. RESULTS: NFV sensitized both, PDAC only and PDAC cocultured with PSC (PDAC: Panc-1, MiaPaCa-2, PSN-1). In Panc-1 and PSN-1 this effect was larger +PSC compared to -PSC. Human pancreatic stellate cells (hPSC) were also sensitized by NFV which reduced p-FAK levels in hPSC, an effect that we previously found to sensitize specifically PDAC/PSC coculture. Contrarily, LY294002 reduced p-Akt in PSC (hPSC and LTC-14) but had no impact on PSC radiation survival. In vitro, nelfinavir sensitized Panc-1 and PSN-1 under normoxic and hypoxic conditions. In PSN-1 xenografts, +PSC led to faster tumor regrowth after radiation vs -PSC. The regrowth delay effect of nelfinavir after radiation was dramatically larger +PSC vs -PSC (time to reach 250mm(3) 183% vs 22%). CONCLUSION: NFV mediated radiosensitization in PDAC with stroma is partly mediated by p-FAK inhibition (Chen et al., 2013). In vitro, NFV sensitizes both normoxic and hypoxic PDAC +/- PSC to a roughly similar extent. The dramatic increased effect of xenograft regrowth inhibition by nelfinavir in tumors with PSC is attributed to vascular normalization (Brunner et al., 2014) rather than direct modification of hypoxia as shown by the tumor regrowth after gemcitabine with NFV.
Assuntos
Inibidores da Protease de HIV/farmacologia , Nelfinavir/farmacologia , Neoplasias Pancreáticas/radioterapia , Células Estreladas do Pâncreas/efeitos dos fármacos , Radiossensibilizantes/farmacologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Nus , Neoplasias Pancreáticas/patologiaRESUMO
We examined the prognostic value of programmed cell death-1 (PD-1) and its ligand (PD-L1) together with CD8+ tumor-infiltrating lymphocytes (TILs) and FOXP3+ Tregs in resectable pancreatic ductal adenocarcinoma (PDAC) samples treated with adjuvant chemotherapy. Whole-mount FFPE tissue sections from 145 pancreatectomies were immunohistochemically stained for PD-1, PD-L1, CD8 and FOXP3. Their expression was correlated with clinicopathological characteristics, and overall survival (OS), progression-free survival (PFS), local progression-free survival (LPFS) and distant metastases free-survival (DMFS), in the context of stroma density (haematoxylin-eosin) and activity (alpha-smooth muscle actin) and in regard to intratumoral lymphoid aggregates. The median OS was 21 months after a mean follow-up of 20 months (range, 2-69 months). In multivariate analysis, high PD-1+ TILs expression was associated with better OS (p = 0.049), LPFS (p = 0.017) and DMFS (p = 0.021). Similar findings were observed for CD8+ TILs, whereas FOXP3 and PD-L1 lacked prognostic significance. Although TIL distribution was heterogeneous, tumors of high stroma density had higher infiltration of CD8+ TILs than loose density stroma and vice versa (p < 0.001), whereas no correlation was found with stromal activity. Sixty (41.4%) tumors contained lymphoid aggregates and the presence of PD-1+ TILs was associated with better OS (p = 0.030), LPFS (p = 0.025) and DMFS (p = 0.033), whereas CD8+ TILs only correlated with superior LPFS (p = 0.039). PD-1+ and CD8+ TILs constitute independent prognostic markers in patients with PDAC treated with adjuvant chemotherapy. Our study provides important insight on the role of PD-1/PD-L1 in the context of desmoplastic stroma and could help guide future immunotherapies in PDAC.
Assuntos
Antígeno B7-H1/metabolismo , Biomarcadores Tumorais/metabolismo , Antígenos CD8/metabolismo , Carcinoma Ductal Pancreático/diagnóstico , Fatores de Transcrição Forkhead/metabolismo , Neoplasias Pancreáticas/diagnóstico , Receptor de Morte Celular Programada 1/metabolismo , Células Estromais/metabolismo , Adolescente , Adulto , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/mortalidade , Carcinoma Ductal Pancreático/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Valor Preditivo dos Testes , Prognóstico , Células Estromais/patologia , Análise de Sobrevida , Adulto JovemRESUMO
Molecular biomarkers are currently evaluated in preclinical and clinical studies in order to establish predictors for treatment decisions in radiation oncology. The receptor tyrosine kinases (RTK) are described in the following text. Among them, the most data are available for the epidermal growth factor receptor (EGFR) that plays a major role for prognosis of patients after radiotherapy, but seems also to be involved in mechanisms of radioresistance, specifically in repopulation of tumour cells between radiotherapy fractions. Monoclonal antibodies against the EGFR improve locoregional tumour control and survival when applied during radiotherapy, however, the effects are heterogeneous and biomarkers for patient selection are warranted. Also other RTK´s such as c-Met and IGF-1R seem to play important roles in tumour radioresistance. Beside the potential to select patients for molecular targeting approaches combined with radiotherapy, studies are also needed to evluate radiotherapy adaptation approaches for selected patients, i.e. adaptation of radiation dose, or, more sophisticated, of target volumes.
Assuntos
Receptores ErbB/metabolismo , Neoplasias/terapia , Medicina de Precisão/métodos , Radioterapia (Especialidade)/métodos , Receptores Proteína Tirosina Quinases/metabolismo , Anticorpos Monoclonais/uso terapêutico , Quimiorradioterapia , Receptores ErbB/antagonistas & inibidores , Humanos , Neoplasias/metabolismo , Inibidores de Proteínas Quinases/uso terapêutico , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Análise de SobrevidaRESUMO
BACKGROUND AND PURPOSE: Nelfinavir can enhance intrinsic radiosensitivity, reduce hypoxia and improve vascularity. We conducted a phase II trial combining nelfinavir with chemoradiotherapy (CRT) for locally advanced inoperable pancreatic cancer (LAPC). MATERIALS AND METHODS: Radiotherapy (50.4Gy/28 fractions; boost to 59.4Gy/33 fractions) was administered with weekly gemcitabine and cisplatin. Nelfinavir started 3-10days before and was continued during CRT. The primary end-point was 1-year overall survival (OS). Secondary end-points included histological downstaging, radiological response, 1-year progression free survival (PFS), overall survival (OS) and treatment toxicity. An imaging sub-study (n=6) evaluated hypoxia ((18)F-Fluoromisonidazole-PET) and perfusion (perfusion CT) during induction nelfinavir. RESULTS: The study closed after recruiting 23 patients, due to non-availability of Nelfinavir in Europe. The 1-year OS was 73.4% (90% CI: 54.5-85.5%) and median OS was 17.4months (90% CI: 12.8-18.8). The 1-year PFS was 21.8% (90% CI: 8.9-38.3%) and median PFS was 5.5months (90% CI: 4.1-8.3). All patients experienced Grade 3/4 toxicity, but many were asymptomatic laboratory abnormalities. Four of 6 patients on the imaging sub-study demonstrated reduced hypoxia and increased perfusion post-nelfinavir. CONCLUSIONS: CRT combined with nelfinavir showed acceptable toxicity and promising survival in pancreatic cancer.
Assuntos
Quimiorradioterapia/métodos , Inibidores da Protease de HIV/uso terapêutico , Nelfinavir/uso terapêutico , Neoplasias Pancreáticas/terapia , Terapia Combinada , Intervalo Livre de Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neoplasias Pancreáticas/patologiaRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is characterized by an abundant desmoplastic stroma. We examined the prognostic value of stroma density and activity in patients with resectable PDAC treated with surgery and adjuvant gemcitabine-based chemotherapy. FFPE-tissue from the pancreatectomy of 145 patients was immunohistochemically stained for haematoxylin-eosin and Masson's trichrome to assess stroma density, and alpha-smooth muscle actin (αSMA) expression for activated pancreatic stellate cells. Their expression was correlated with clinicopathological characteristics as well as overall survival (OS), progression-free survival (PFS), local progression-free survival (LPFS) and distant metastases free-survival (DMFS). After a mean follow-up of 20 months (range, 2-69 months), the median OS was 21 months and the 3-year OS was 35.7%. In multivariate analysis, highly-dense stroma was an independent prognostic parameter for OS (p = 0.001), PFS (p = 0.007), LPFS (p = 0.001) and DMFS (p = 0.002), while αSMA expression lacked significance. Interestingly, highly-dense stroma retained significance for the four clinical endpoints only in early (pT1-2) but not late (pT3-4) stage tumors. Additionally, late pT-stage (pT3-4), the presence of lymph node metastases (pN+ vs pN0), perineural/neural invasion and administration of adjuvant chemotherapy also correlated with prognosis in multivariate analysis. Altogether, stroma density constitutes an independent prognostic marker in PDAC patients treated with adjuvant chemotherapy. Our findings highlight the dynamic complexity of desmoplasia and indicate that highly-dense stroma is correlated with better outcome. Further validation of the prognostic value of stroma as a biomarker and its role in PDAC patients after adjuvant chemotherapy is warranted and will be performed in a prospective study.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Ductal Pancreático/secundário , Pancreatectomia/mortalidade , Neoplasias Pancreáticas/patologia , Células Estromais/patologia , Idoso , Carcinoma Ductal Pancreático/terapia , Terapia Combinada , Feminino , Seguimentos , Humanos , Metástase Linfática , Masculino , Gradação de Tumores , Invasividade Neoplásica , Estadiamento de Neoplasias , Neoplasias Pancreáticas/terapia , Prognóstico , Taxa de Sobrevida , Neoplasias PancreáticasRESUMO
Colony formation is the gold standard assay for determining reproductive cell death after radiation treatment, since effects on proliferation often do not reflect survival. We have developed a high-throughput radiosensitivity screening method based on clonogenicity and screened a siRNA library against kinases. Thiamine pyrophosphokinase-1 (TPK1), a key component of Vitamin B1/thiamine metabolism, was identified as a target for radiosensitization. TPK1 knockdown caused significant radiosensitization in cancer but not normal tissue cell lines. Other means of blocking this pathway, knockdown of thiamine transporter-1 (THTR1) or treatment with the thiamine analogue pyrithiamine hydrobromide (PyrH) caused significant tumor specific radiosensitization. There was persistent DNA damage in cells irradiated after TPK1 and THTR1 knockdown or PyrH treatment. Thus this screen allowed the identification of thiamine metabolism as a novel radiosensitization target that affects DNA repair. Short-term modulation of thiamine metabolism could be a clinically exploitable strategy to achieve tumor specific radiosensitization.
Assuntos
Neoplasias/metabolismo , Neoplasias/radioterapia , Tiamina/metabolismo , Linhagem Celular Tumoral , Ensaio de Unidades Formadoras de Colônias , Dano ao DNA , Células HCT116 , Células HeLa , Ensaios de Triagem em Larga Escala , Humanos , Piritiamina/farmacologia , Tolerância a Radiação , Radiossensibilizantes/farmacologia , Tiamina Pirofosfoquinase/metabolismo , TransfecçãoRESUMO
Combining chemotherapy with radiotherapy has resulted in significant clinical improvements in many different tumour types. However, the non-specific mechanisms by which these drugs exert their effects mean that this is often at the expense of increased side effects. Previous attempts at using targeted drugs to induce more tumour specific radiosensitisation have been generally disappointing. Although cetuximab, an EGFR monoclonal antibody, resulted in improved overall survival in HNSCC when combined with radiotherapy, it has failed to show benefit when added to chemo-radiotherapy. In addition, our inability to successfully use drug treatments to reverse tumour hypoxia is underlined by the fact that no such treatment is currently in widespread clinical use. The reasons for these failures include the lack of robust biomarkers, and the previous use of drugs with unacceptable side-effect profiles. Despite these disappointments, there is reason for optimism. Our improved understanding of key signal transduction pathways and of tumour specific DNA repair deficiencies has produced new opportunities to specifically radiosensitise tumours. Novel strategies to reduce tumour hypoxia include the use of drugs that cause vascular normalisation and drugs that reduce tumour oxygen consumption. These new strategies, combined with better compounds at our disposal, and an ability to learn from our previous mistakes, mean that there is great promise for future drug-radiotherapy combinations to result in significant clinical benefits.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Quimiorradioterapia , Reparo do DNA/efeitos dos fármacos , Imunoterapia , Neoplasias/metabolismo , Neoplasias/terapia , Radiossensibilizantes/administração & dosagem , Radiossensibilizantes/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Hipóxia Celular/efeitos dos fármacos , Quimiorradioterapia/efeitos adversos , Quimiorradioterapia/métodos , Quimiorradioterapia Adjuvante/efeitos adversos , Ensaios Clínicos como Assunto , Terapia Combinada/métodos , Humanos , Terapia de Alvo Molecular/métodos , Neoplasias/genética , Neoplasias/fisiopatologia , Transdução de Sinais/efeitos dos fármacos , Falha de TratamentoRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death. Despite improvements in the clinical management, the prognosis of PDAC remains dismal. In the present comprehensive review, we will examine the knowledge of PDAC genetics and the new insights into human genome sequencing and clonal evolution. Additionally, the biology and the role of the stroma in tumour progression and response to treatment will be presented. Furthermore, we will describe the evidence on tumour chemoresistance and radioresistance and will provide an overview on the recent advances in PDAC metabolism and circulating tumour cells. Next, we will explore the characteristics and merits of the different mouse models of PDAC. The inflammatory milieu and the immunosuppressive microenvironment mediate tumour initiation and treatment failure. Hence, we will also review the inflammatory and immune escaping mechanisms and the new immunotherapies tested in PDAC. A better understanding of the different mechanisms of tumour formation and progression will help us to identify the best targets for testing in future clinical studies of PDAC.
Assuntos
Carcinoma Ductal Pancreático , Imunoterapia , Neoplasias Pancreáticas , Tolerância a Radiação , Animais , Vacinas Anticâncer/uso terapêutico , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/imunologia , Carcinoma Ductal Pancreático/terapia , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/genética , Resistencia a Medicamentos Antineoplásicos/imunologia , Humanos , Imunoterapia/métodos , Inflamação/patologia , Camundongos , Células Neoplásicas Circulantes/imunologia , Células Neoplásicas Circulantes/patologia , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/terapia , Tolerância a Radiação/genética , Neoplasias PancreáticasRESUMO
The ataxia telangiectasia and Rad3-related (ATR) plays an important role in maintaining genome integrity during DNA replication through the phosphorylation and activation of Chk1 and regulation of the DNA damage response. Preclinical studies have shown that disruption of ATR pathway can exacerbate the levels of replication stress in oncogene-driven murine tumors to promote cell killing. Additionally, inhibition of ATR can sensitise tumor cells to radiation or chemotherapy. Accumulating evidence suggests that targeting ATR can selectively sensitize cancer cells but not normal cells to DNA damage. Furthermore, in hypoxic conditions, ATR blockade results in overloading replication stress and DNA damage response causing cell death. Despite the attractiveness of ATR inhibition in the treatment of cancer, specific ATR inhibitors have remained elusive. In the last two years however, selective ATR inhibitors suitable for in vitro and - most recently - in vivo studies have been identified. In this article, we will review the literature on ATR function, its role in DDR and the potential of ATR inhibition to enhance the efficacy of radiation and chemotherapy.