Introduction the objective was to evaluate the impact of IDH1 R132H mutation, MGMT methylation and PD-L1 expression in high grade glioma that received standard therapy (surgery, radiation and chemotherapy) to overall survival (OS). Methods this is a retrospective study of 35 high grade glioma cases. Genotyping of IDH1 gene alteration on the mutation hotspot R132 (Sanger sequencing method with Applied Biosystems 3500 Genetic Analyzer), EZ DNA Methylation-Gold kit (Zymo Research) is used to study the methylation, Cell line BT549 (ATCC HTB-122) and HCT-116 (ATCC CCL-247) were used as unmethylated control and partially methylated control respectively. Anti-human PD-L1 antibody clone E1L3N ® from Cell Signalling Technology (USA) and Rabbit XP ® were used to see PDL-1 expression. Results anaplastic astrocytoma cases had more MGMT promoter methylation (50%) than glioblastoma multiforme (GBM) (20%), more IDH1 R132H mutation (42%) than GBM (4.3%). Immunohistochemistry tumor proportion score method (TPS) identified 17% and 8.7% were PD-L1 positive in AA and GBM groups, respectively. Cases with IDH1 R132H mutation and MGMT methylation still showed better OS although with high PD-L1 expression. Conclusion IDH1 R132H mutation and MGMT methylation were good prognostic markers. High expression of PD-L1 apparently might not indicate poor overall survival in the presence of IDH1 R132 mutation and MGMT methylation.
Background Lung cancer patients with mutations in epidermal growth factor receptor (EGFR) gene are treated with tyrosine kinase inhibitor (TKI). Aims We aimed to evaluate polymerase chain reaction (PCR)–high‐resolution melting (HRM), restriction fragment length polymorphism (RFLP), and direct sequencing (DS) to detect EGFR mutations in cell‐free DNA (cfDNA) before and after TKI treatment in real‐world settings of a developing country. Methods Paired cytology and plasma samples were collected from 116 treatment‐naïve lung cancer patients. DNA from both plasma and cytology specimens was isolated and analyzed using PCR‐HRM (to detect exon 19 insertion/deletion), RFLP (to genotypes L858R and L861Q), and DS (to detect uncommon mutations G719A, G719C, or G719S [G719Xaa] in exon 18 and T790M and insertion mutations in exon 20). Results EGFR genotypes were obtained in all 116 (100%) cfDNA and 110/116 (94.82%) of cytological specimens of treatment‐naïve patient (baseline samples). EGFR‐activating mutations were detected in 46/110 (40.6%) plasma samples, and 69/110 (63.2%) mutations were found in routine cytology samples. Using cytological EGFR genotypes as reference, we found that sensitivity and specificity of baseline plasma EGFR testing varied from 9.1% to 39.39% and 83.12% to 96.55%, respectively. In particular, the sensitivity and specificity of this assay in detecting baseline T790M mutations in exon 20 were 30% and 89.58%, respectively. Three months after TKI treatment, plasma T790M and insertion exon 20 mutations appeared in 5.4% and 2.7% patients, respectively. Conclusions Despite low sensitivity, combined DS, RFLP, and PCR‐HRM was able to detect EGFR mutations in plasma cfDNA with high specificity. Moreover, TKI resistance exon 20 insertions mutation was detected as early as 3 months post TKI treatment.
Background: Indonesia has the highest cigarette consumption in the world. We explored the clinical impact of smoking on the prevalence of EGFR and K-RAS mutations and survival in this prospective study. Methods: 143 treatment naive lung cancer patients were recruited from Persahabatan Hospital, a national tertiary hospital. DNA from cytological specimens had been extracted and genotyped for both EGFR and K-RAS mutations using a combination of PCR high resolution melting, restriction fragment length polymorphism (RFLP) and direct DNA sequencing. Results: EGFR mutation frequency in never smokers (NS) and ever smokers (ES) were 75% and 56% ( p = 0.0401), respectively. In this cohort, the overall K-RAS mutation rate was 7%. Neither gender nor smoking history were associated with K-RAS mutation significantly. However, K-RAS transversion mutations were more common in male ES than transition mutations. Smoking history did not affect EGFR and K-RAS mutation frequencies in women. Concurrent EGFR / K-RAS mutation rate was 2.8% (4 of 143 patients). Four out of 91 EGFR mutation positive patients (4.4%) had simultaneous K-RAS mutation. Conclusions: In region where cigarette consumption is prevalent, smoking history affected frequencies of EGFR and K-RAS mutations, mainly in males.
clinical course has been reported to occur during pregnancy. Pregnancy-associated plasma protein-A (PAPP-A) is a metalloproteinase that cleaves insulin-like growth factor 1 (IGF-1) from a circulating complex formed with insulin-like growth factor binding protein 4 (IGFBP4). Increased bioavailability of IGF1 subsequently impacts on tumour biology. PAPP-A is produced in increasing levels until term by the syncytiotrophoblast cells in the human placenta. High levels of PAPP-A have been associated with progression in other tumour types. Methods: Eleven breast cancer cell lines were examined to characterise the components of the PAPP-A/IGF axis in breast cancer. PAPP-A mRNA expression was analysed by quantitative RT-PCR. The effect of PAPP-A expression on cell migration in the MDA-MB-453 cell line was subsequently evaluated by using neutralising antibodies. An analysis of The Cancer Genome Atlas (TCGA) publicly available transcriptome profiling dataset was also performed, through specific query of expression clustering. Results: PAPP-A expression was noted in three of the cell lines. In the MDA-MB-453 cell line, which expressed high levels of PAPP-A, neutralising antibodies to PAPP-A and IGFBP4 significantly reduced migratory capabilities. Analysis of the TCGA dataset revealed that alteration (mainly upregulation) in PAPP-A expression resulted in worse overall survival (n ¼ 1104, p ¼ 0.13). Median survival with alteration in PAPP-A expression was 74 months compared with 113 months in patients without alteration in PAPP-A. Conclusions: Our data suggest that PAPP-A plays a role in the progression of breast cancer, which may be particularly relevant in pregnancy. The insights gained from this research open up the possibility of indirect and specific therapeutic targeting of IGF to halt the progression of breast cancer.
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