Antiproliferative efficacies but minor drug transporter inducing effects of paclitaxel, cisplatin, or 5-fluorouracil in a murine xenograft model for head and neck squamous cell carcinoma

Drug-induced multidrug resistance (MDR) has been linked to overexpression of drug transporting proteins in head and neck squamous cell carcinoma (HNSCC) in vitro. The aim of this work was to reassess these findings in a murine xenograft model. NOD-SCID mice xenotransplanted with 106 HNO97 cells were treated for four consecutive weeks with weekly paclitaxel, biweekly cisplatin (both intraperitoneal), or 5-fluorouracil (5-FU, administered by osmotic pump). Tumor volume and body weight were weekly documented. Expression of drug transporters and Ki-67 marker were examined using quantitative real-time polymerase chain reaction and/or immunohistochemistry. Both paclitaxel and cisplatin significantly reduced tumor volumes after 2–3 weeks. 5-FU-treated animals had significantly lower body weights after 2 or 4 weeks of chemotherapy. None of the drugs affected expression of drug transporters at the mRNA level. However, P-glycoprotein (Pgp) protein expression was increased by paclitaxel (P < 0.01). Ki-67 expression did not change during treatment irrespective of the drug applied. Paclitaxel and cisplatin are effectively tumor volume reducing drugs in a murine xenograft model of HNSCC. Paclitaxel enhanced Pgp expression at the protein level, but not at the mRNA level suggesting transcriptional induction to be of minor relevance. In contrast, posttranscriptional mechanisms or Darwinian selection of intrinsically drug transporter overexpressing MDR cells might lead to iatrogenic chemotherapy resistance in HNSCC.Fil: Theile, Dirk. Universität Heidelberg; AlemaniaFil: Gal, Zoltan. Universität Heidelberg; AlemaniaFil: Warta, Rolf. Universität Heidelberg; AlemaniaFil: Rigalli, Juan Pablo. Universität Heidelberg; Alemania. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lahrmann, Bernd. Universität Heidelberg; AlemaniaFil: Grabe, Niels. Universität Heidelberg; AlemaniaFil: Herold Mende, Christel. Universität Heidelberg; AlemaniaFil: Dyckhoff, Gerhard. Universität Heidelberg; AlemaniaFil: Weiss, Johanna. Universität Heidelberg; Alemani

Spatial transcriptome analysis reveals Notch pathway-associated prognostic markers in IDH1 wild-type glioblastoma involving the subventricular zone

Background: The spatial relationship of glioblastoma (GBM) to the subventricular zone (SVZ) is associated with inferior patient survival. However, the underlying molecular phenotype is largely unknown. We interrogated an SVZ-dependent transcriptome and potential location-specific prognostic markers. Methods: mRNA microarray data of a discovery set (n = 36 GBMs) were analyzed for SVZ-dependent gene expression and process networks using the MetaCore™ workflow. Differential gene expression was confirmed by qPCR in a validation set of 142 IDH1 wild-type GBMs that was also used for survival analysis. Results: Microarray analysis revealed a transcriptome distinctive of SVZ+ GBM that was enriched for genes associated with Notch signaling. No overlap was found to The Cancer Genome Atlas’s molecular subtypes. Independent validation of SVZ-dependent expression confirmed four genes with simultaneous prognostic impact: overexpression of HES4 (p = 0.034; HR 1.55) and DLL3 (p = 0.017; HR 1.61) predicted inferior, and overexpression of NTRK2 (p = 0.049; HR 0.66) and PIR (p = 0.025; HR 0.62) superior overall survival (OS). Additionally, overexpression of DLL3 was predictive of shorter progression-free survival (PFS) (p = 0.043; HR 1.64). Multivariate analysis revealed overexpression of HES4 to be independently associated with inferior OS (p = 0.033; HR 2.03), and overexpression of DLL3 with inferior PFS (p = 0.046; HR 1.65). Conclusions: We identified four genes with SVZ-dependent expression and prognostic significance, among those HES4 and DLL3 as part of Notch signaling, suggesting further evaluation of location-tailored targeted therapies

Human Leucocyte Antigens as Prognostic Markers in Head and Neck Squamous Cell Carcinoma

Background: The induction and regulation of immune responses depend on human leucocyte antigen (HLA) molecules that present peptides derived from mutated neoantigens or tumor-associated antigens to cytotoxic T cells. The natural variation of HLA molecules might differ between tumor patients and the normal population. Thus, there might be associations between the frequencies of HLA alleles and the survival of tumor patients. Methods: This issue was studied in a cohort of 84 patients with head and neck squamous cell carcinomas (HNSCCs) of different localizations. The cohort was followed up for more than 10 years. HLA-A/B/C CTS-PCR-SSP typing at 1 field level from blood samples was performed, and the results were correlated with survival. Results: HLA-A*02 was the most prevalent allele in our cohort and was present in 51.1% of patients. The HLA-A*25 and HLA-C*06 alleles exhibited a significantly higher frequency in cancer patients than in the normal population of 174 blood and kidney donors (p = 0.02 and p = 0.01, respectively, Fisher&rsquo;s exact test). For HLA-C*04, a negative impact on overall survival in univariate analysis (p = 0.045) and a negative, but statistically insignificant effect on survival toward poorer survival in multivariate analysis (HR: 1.82; 95% CI: 0.99&ndash;3.34, p = 0.053) were observed. In addition, HLA-A*02 was also beneficial for overall survival and progression-free survival in multivariate analysis (HR 0.54; 95% CI: 0.31&ndash;0.92; p = 0.023). Conclusion: HLA-A*02 allele expression might not only predict better survival but might also indicate superior tumor antigen presentation and, thus, help to select patients who could benefit from T-cell-dependent immunotherapies

Chemoradiotherapy but Not Radiotherapy Alone for Larynx Preservation in T3. Considerations from a German Observational Cohort Study

For advanced laryngeal cancers, after randomized prospective larynx preservation studies, nonsurgical therapy has been applied on a large scale as an alternative to laryngectomy. For T4 laryngeal cancer, poorer survival has been reported after nonsurgical treatment. Is there a need to fear worse survival also in T3 tumors? The outcomes of 121 T3 cancers treated with pCRT, pRT alone, or surgery were evaluated in an observational cohort study in Germany. In a multivariate Cox regression of the T3 subgroup, no survival difference was noted between pCRT and total laryngectomy with risk-adopted adjuvant (chemo)radiotherapy (TL ± a(C)RT) (HR 1.20; 95%-CI: 0.57–2.53; p = 0.63). However, survival was significantly worse after pRT alone than after TL ± a(C)RT (HR 4.40; 95%-CI: 1.72–11.28, p = 0.002). A literature search shows that in cases of unfavorable prognostic markers (bulky tumors of 6–12 ccm, vocal cord fixation, minimal cartilage infiltration, or N2–3), pCRT instead of pRT is indicated. In cases of pretreatment dysphagia or aspiration requiring a feeding tube or tracheostomy, gross or multiple cartilage infiltration, or tumor volume &gt; 12 ccm, outcomes after pCRT were significantly worse than those after TL. In these cases, and in cases where pCRT is indicated but the patient is not suitable for the addition of chemotherapy, upfront total laryngectomy with stage-appropriate aRT is recommended even in T3 laryngeal cancers

Could Primary Chemoradiotherapy in T2 Glottic Cancers Yield Results Comparable to Primary Radiotherapy in T1? Considerations from 531 German Early Stage Patients

T1 glottic cancer is a highly treatable disease with local control (LC) rates over 90% by either primary radiotherapy (pRT) or transoral laser microsurgery (TLM). LC of T2 glottic cancers is 15 percent points poorer on average. However, salvage after pRT entails more than 50% total laryngectomy. Therefore, there is a need for enhanced LC. Altered fractionation regimens improved LC in T1 but not in T2. For this reason, for T2, alternative strategies must be considered. In a large observational cohort study including 531 early-stage laryngeal cancers, a small number of patients were treated with primary chemoradiotherapy (pCRT). In multivariable analysis, factors associated with significantly poorer outcomes included age, comorbidities, supraglottic localization, and T category. While there was a significant difference between pRT and surgery (HR 1.79; 95%-CI: 1.15–2.79), there was none between pCRT and surgery (HR 0.70; 95%-CI: 0.33–1.51). There is evidence from the literature that pCRT in early glottic cancers could yield results that surpass the limits so far experienced in radiotherapy alone with acceptable toxicity. Thus, prospective randomized studies with larger numbers of patients are warranted

An Observational Cohort Study on 194 Supraglottic Cancer Patients: Implications for Laser Surgery and Adjuvant Treatment

Supraglottic laryngeal cancer is characterized by poor prognosis. In contrast, excellent outcomes have been published in early-stage supraglottic cancers after laser surgery in single-institutional series in centers of excellence. Are these results reproducible in the normal clinical practice of less specialized facilities? As part of an observational cohort study, the outcomes of 194 supraglottic cancer patients were assessed after treatment by larynx-preserving surgery (transoral laser microsurgery [TLM] or open partial laryngectomy [OPL]) or total laryngectomy (TL), with each having risk-adopted adjuvant treatment, or primary (chemo-)radiotherapy (pCRT or pRT). In early-stage supraglottic cancers, TLM achieved a 5-year overall survival (5-year OS) of 62.0%. No significant survival difference could be discerned between patients with and without adjuvant treatment (HR 1.47; 95% CI: 0.80 2.69). The comparison between pCRT and pRT patients suggests that CRT is more effective in supraglottic cancer. The 5-year OS rate achieved in our multiinstitutional setting is comparable to that reached in laser surgery centers of excellence (59.4–76.0%). According to our data and supported by the literature, adjuvant RT (aRT) is not sufficiently effective in supraglottic cancers. In case adjuvant therapy is indicated, adjuvant chemoradiation (aCRT) could be recommended

Identification of a Prognostic Hypoxia-Associated Gene Set in IDH-Mutant Glioma

Glioma growth is often accompanied by a hypoxic microenvironment favorable for the induction and maintenance of the glioma stem cell (GSC) phenotype. Due to the paucity of cell models of Isocitrate Dehydrogenase 1 mutant (IDH1mut) GSCs, biology under hypoxic conditions has not been sufficiently studied as compared to IDH1 wildtype (IDH1wt) GSCs. We therefore grew well-characterized IDH1mut (n = 4) and IDH1wt (n = 4) GSC lines under normoxic (20%) and hypoxic (1.5%) culture conditions and harvested mRNA after 72 h. Transcriptome analyses were performed and hypoxia regulated genes were further analyzed using the expression and clinical data of the lower grade glioma cohort of The Cancer Genome Atlas (LGG TCGA) in a confirmatory approach and to test for possible survival associations. Results show that global expression changes were more pronounced in IDH1wt than in IDH1mut GSCs. However, when focusing on known hypoxia-regulated gene sets, enrichment analyses showed a comparable regulation in both IDH1mut and IDH1wt GSCs. Of 272 significantly up-regulated genes under hypoxic conditions in IDH1mut GSCs a hypoxia-related survival score (HRS-score) of five genes (LYVE1, FAM162A, WNT6, OTP, PLOD1) was identified by the Least Absolute Shrinkage and Selection Operator (LASSO) algorithm which was able to predict survival independent of age, 1p19q co-deletion status and WHO grade (II vs. III) in the LGG TCGA cohort and in the Rembrandt dataset. Altogether, we were able to identify and validate a novel hypoxia-related survival score in IDH1mut GSCs consisting of five hypoxia-regulated genes which was significantly associated with patient survival independent of known prognostic confounders

Large-Scale Drug Screening in Patient-Derived IDHmut Glioma Stem Cells Identifies Several Efficient Drugs among FDA-Approved Antineoplastic Agents

The discovery of the isocitrate dehydrogenase (IDH) mutation in glioma led to a paradigm shift on how we see glioma biology. Difficulties in cultivating IDH mutant glioma stem cells (IDHmut GSCs) resulted in a paucity of preclinical models in IDHmut glioma, limiting the discovery of new effective chemotherapeutic agents. To fill this gap, we used six recently developed patient-derived IDHmut GSC lines and performed a large-scale drug screening with 147 Food and Drug Administration (FDA)-approved anticancer drugs. GSCs were subjected to the test compounds for 72 h in concentrations ranging from 0.0001 to 1 &micro;M. Cell viability was assessed by CellTiterGlo and the induction of apoptosis by flow cytometry with Annexin V/propidium iodide staining. The initial screen was performed with two IDHmut GSC lines and identified seven drugs (bortezomib, carfilzomib, daunorubicin, doxorubicin, epirubicin, omacetaxine, plicamycin) with a substantial antiproliferative activity, as reflected by half maximal inhibitory concentrations (IC50) below 1 &micro;M and maximum inhibitory effects (Emax) below 25%. These findings were validated in an additional four IDHmut GSC lines. The candidate drugs, of which plicamycin and omacetaxine are known to cross the blood brain barrier, were used for subsequent cell death analyses. A significant induction of apoptosis was observed at IC50 values of the respective drugs. In summary, we were able to identify seven FDA-approved drugs that should be further taken into clinical investigations for the treatment of IDHmut gliomas

Extent of Resection, MGMT Promoter Methylation Status and Tumor Location Independently Predict Progression-Free Survival in Adult Sporadic Pilocytic Astrocytoma

In adults, pilocytic astrocytomas (PA) account for less than 2% of gliomas, resulting in uncertainty regarding the clinical course and optimal treatment, particularly in cases where gross total resection (GTR) could not be achieved. Moreover, information on molecular markers and their prognostic impact is sparse. In order to improve risk stratification, we analyzed our institutional series of 58 patients aged 17 years and older with histology-proven intracranial PA World Health Organization grade I for clinical and molecular prognosticators. Anaplastic and NF1-associated tumors were excluded. O-6-methylguanine-DNA methyltransferase (MGMT) promoter methylation status was determined by pyrosequencing or 450k/850k DNA methylation array. A univariate log-rank test and multivariate StepAIC were applied to identify prognostic factors. The median age was 30 years (range 17&ndash;66). Tumors were located in the cerebral/cerebellar hemispheres, midline structures and cerebello-pontine angle in 53%, 38% and 9%. MGMT promoter methylation was present in eight patients (14%). GTR (39/58 patients) significantly reduced the likelihood of tumor recurrence (p = 0.0001). Tumor relapse occurred in 16 patients (28%) after a median progression-free survival (PFS) of 135 months (range 6&ndash;153 months); there was one tumor-related death. PFS at 5 and 10 years was 67% and 53%. In multivariate analysis, PFS was significantly prolonged in patients with GTR (HR 0.1; CI 0.03&ndash;0.37; p &lt; 0.001), unmethylated MGMT promoter (HR 0.18; CI 0.05&ndash;0.64; p = 0.009) and midline tumors (HR 0.21; CI 0.06&ndash;0.78; p = 0.02). In conclusion, MGMT promoter methylation status and tumor location were identified as novel prognostic factors in adult PAs, pointing at distinct molecular subtypes and detecting patients in need of close observance and intensified treatment