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Anant Mohan
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EP1.01 - Advanced NSCLC (ID 150)
- Event: WCLC 2019
- Type: E-Poster Viewing in the Exhibit Hall
- Track: Advanced NSCLC
- Presentations: 1
- Now Available
- Moderators:
- Coordinates: 9/08/2019, 08:00 - 18:00, Exhibit Hall
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EP1.01-22 - Liquid Biopsy in the Detection of EGFR Activating and Resistance Mutations in Advanced Pulmonary Adenocarcinoma (Now Available) (ID 1761)
08:00 - 18:00 | Author(s): Anant Mohan
- Abstract
Background
Approximately 30% of the advanced pulmonary adenocarcinoma (PADC) patients are found to be mutated with epidermal growth factor receptor (EGFR). Nearly 60% of EGFR mutated patients develop T790M resistance mutation against which third generation TKIs are available. Detection of T790M mutation is a challenge due to non availability of enough tumour tissues in patients receiving first-generation EGFR-TKI treatment. Liquid biopsies (cell- free DNA) are promising for detection of EGFR activating and resistance mutations. The current study focuses on detection of EGFR mutations in cell-free DNA from patients with histologically confirmed EGFR mutation status.
Method
50 PADC patients (22 males; 28 females with mean age 63 years) were studied for EGFR mutations. Out of them 40 were TKI naïve and 10 patients were treated with EGFR-TKIs. Follow up biopsy from TKI treated patients was not available for resistant mutation studies. Real-time PCR and Digital droplet PCR techniques were adopted for studying EGFR exon 19 (deletions), 20 (T790M) and 21 (L858R) mutations in tissue and plasma samples respectively.
Result
Clinically all patients were in stage III-IV and 52% were never smokers. Out of 40 TKI naive cases, 20 cases were positive for EGFR activating mutations in tissue and 17 in plasma. Twenty cases were negative for EGFR mutations in both tissue and plasma. In 10 EGFR-TKI treated patients, founder activating mutation was still present in plasma of 7 cases and 3 of them also showed T790M resistance mutation. Two patients carrying this resistant mutation died subsequently. Remaining three TKI treated patients did not harbour even founder activating mutation in the plasma.
Conclusion
We observed a concordance of 88% (44/50) between liquid biopsy and tumour tissue. Our study highlights role of liquid biopsy as an alternative for detecting EGFR activating and resistance mutations in advanced PADC. EGFR mutation analysis in liquid biopsy can be helpful in those patients where the tissue biopsy is not possible.
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EP1.03 - Biology (ID 193)
- Event: WCLC 2019
- Type: E-Poster Viewing in the Exhibit Hall
- Track: Biology
- Presentations: 1
- Now Available
- Moderators:
- Coordinates: 9/08/2019, 08:00 - 18:00, Exhibit Hall
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EP1.03-20 - Diagnostic and Prognostic Utility of Differentially Expressed Circulating MicroRNAs in Indian NSCLC Patients (Now Available) (ID 875)
08:00 - 18:00 | Author(s): Anant Mohan
- Abstract
Background
The presence of circulating tumour DNA, mRNA and non-coding RNAs, such as microRNA (miRNA), in the serum and plasma of cancer patients has sparked great interest because conventional diagnostic tests tend to be imperfect and more invasive, posing logistic difficulties for serial tumour sampling. Hence, identification of differentially expressed circulating miRNAs in the serum of non-small cell lung cancer (NSCLC) patients may have potential as cancer biomarkers for diagnosis, prognosis and predicting therapeutic response.
Method
For the identification of differentially expressed miRNAs in the serum of NSCLC patients, we performed small RNA sequencing with illumina HiSeq 2000 platform (n=10; 4 NSCLC patients and 6 controls). The expression profile of miRNAs in each subject was analyzed using miRNAkey software and fold change was performed to identify differentially expressed miRNAs in NSCLC as compared to controls. We validated the expression of few differentially expressed miRNAs in the serum of 75 NSCLC patients and 40 controls using miScript qRT-PCR assay. The expression of miRNAs was correlated with overall survival (OS), progression-free survival (PFS), response to therapy and various clinico-pathological parameters.
Result
The mean age of NSCLC patients and controls was 56.2 years and 55.3 years, respectively (p = 0.3242). Majority of the NSCLC patient and controls were male. 67% of NSCLC patients and 53% of controls were smokers (p = 0.099). Global miRNA profiling revealed 16 differentially expressed miRNAs (cut-off: fold change > 2.0, or p < 0.05, or both) in the serum of NSCLC patients as compared to controls. Our qRT-PCR data revealed significant down-regulation of miR-15a-5p, miR-320a, miR-25-3p, miR-192-5p, let-7d-5p, let-7e-5p, miR-148a-3p and miR-92a-3p in the serum of NSCLC patients as compared to controls. None of the miRNAs were correlated with survival outcome and therapeutic response. The expression of miR-320a, miR-25-3p and miR-148a-3p significantly correlated with stage, while miR-375 expression significantly correlated with lymph node involvement and pleural effusion.
Conclusion
The expression of majority of miRNAs was down-regulated in the serum of NSCLC patients as compared to controls. Some of the miRNAs, such as miR-375 and miR-320a, are less studied for their involvement in the pathogenesis of NSCLC. Hence, future mechanistic studies are warranted to elucidate their role in disease biology and as candidate biomarkers for diagnosis and prognosis of NSCLC.
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P1.14 - Targeted Therapy (ID 182)
- Event: WCLC 2019
- Type: Poster Viewing in the Exhibit Hall
- Track: Targeted Therapy
- Presentations: 1
- Now Available
- Moderators:
- Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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P1.14-48 - Whole Exome Sequencing (WES) in Non-Small Cell Lung Carcinoma (NSCLC): Identification of Novel Biomarkers (Now Available) (ID 1712)
09:45 - 18:00 | Author(s): Anant Mohan
- Abstract
Background
Significant advancement has been made in the treatment of patients with pulmonary adenocarcinoma (ADC) on the basis of the molecular profile. However, no such molecular target exists for squamous cell carcinoma (SCC) or small cell lung cancer (SCLC). WES has been in wide use for the discovery of new genetic markers, which may offer more information for the development of personalized medicine for all subtypes of lung cancer. The aim of the current study is to find out novel genetic markers for NSCLC which can be used as a universal biomarker for the treatment.
Method
WES of 20 advanced NSCLC patients (10 ADC and 10 SCC) was done on the Illumina HiSeqX10 with paired end 151bp chemistry. The Male: female ratio was 5.6:1 and a median age 56 years. Only 3 patients were non-smoker and in one case smoking history was unknown. 75% of patients were present with co-morbidities, all patients were either in stage III or IV and performance status was either 1 or 2.
Result
Table 1: WES data statistics and the total number of variants identified: S.No WES data Numbers 1 Average raw data 13.6 GB 2 Average processed data 12.1 GB 3 Average reads aligned 80.2% 4 Average total variations 34173 5 Average SNPs 26030 6 Average INDEL 81423 7 Average frameshift deletions 2185 8 Average frameshift insertions 3571 9 Average non-frameshift deletions 368 10 Average non-frameshift insertions 1292 11 Average non-synonymous 13418 12 Average stop gain 10337 13 Average stop loss 38 14 Average synonymous SNPs 11607 15 Average variations reported in databases 20269 16 Unknown mutations 659 WES data statistics has been depicted in table 1 including number of variants identified. After excluding common variants (MAF > 0.05), a total of 23 rare variants (0 < MAF < 0.01), possibly linked to lung carcinoma was identified in the exome. Further stringent filtering showed allele frequencies of mutations in GPRIN2 (G protein-regulated inducer of neurite outgrowth 2), KCNJ18 (potassium voltage-gated channel subfamily J member 18) and TEKT4 (Tektin 4) genes which were present in all the cases. The variations of amino acid substitutions were i) TEKT4: p.A223T and p.A405T (rs75603622); ii) GPRIN2: p.V241M (rs9422022), p.V47M (rs3127819); iii) KCNJ18: p.L211F (rs1435776313), p.I262S (rs1450551937)
Conclusion
Although the mechanism of GPRIN2, KCNJ12 and TEKT4 in tumorigenesis is unclear; our results suggest that these may play a major role in NSCLC and it is worth to be investigated in future. Validation of these genes is under process.
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P2.09 - Pathology (ID 174)
- Event: WCLC 2019
- Type: Poster Viewing in the Exhibit Hall
- Track: Pathology
- Presentations: 1
- Moderators:
- Coordinates: 9/09/2019, 10:15 - 18:15, Exhibit Hall
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P2.09-06 - Epidermal Growth Factor Receptor (EGFR) Mutation Spectrum in Pulmonary Adenocarcinomas: Indian Experience (ID 2447)
10:15 - 18:15 | Author(s): Anant Mohan
- Abstract
Background
Incidence of lung cancer is rising in India, which is the major cause of mortality worldwide. The identification of EGFR mutations and ALK fusions in lung cancer has changed the treatment paradigm. The use of tyrosine kinase inhibitor (TKI) is now well established in clinical practice as it improves treatment outcomes. Molecular testing using tumor tissue is the gold standard for targeted therapies. The aim of the present study is to analyze the exact frequency of EGFR activating, rare and coexisting mutations in pulmonary adenocarcinoma (ADC).
Method
A total of 783 ADC (755 biopsies and 28 cytology aspirate smears) cases were studied. The male: female ratio was 1: 0.3 with age ranging from 20 to 78 years. During grossing, two tumor blocks were made, one for immunohistochemistry studies and other for mutation testing. In high- cellularity, low- tumor fraction cases, tumor enrichment was done by manual microdissection. We used more than one aspirate smears/biopsy blocks in cases with low- cellularity, high- tumor fraction. Probe-based real-time PCR (RT-PCR) technique was used to analyze EGFR hotspots mutations (exons 18 to 21). Before proceeding for molecular testing, histo/cytomorphology, tumour content and DNA quality were determined. For checking the quality of DNA, RT-PCR was done using wild type EGFR exon 2 primers.
Result
About 99% cases passed through the quality check of DNA and only 1.02% (8/783) cases failed. Overall, 234 patients (29.8%) were positive for EGFR mutations. The distribution of different types of EGFR mutations is shown in the Table 1. The common EGFR mutations were Exon 19 del and Exon 21 L858R which were equally distributed between both the genders. The EGFR mutations were more in non-smokers. T790M was present at baseline in around 3% TKI naive cases and it coexisted mostly with Exon 19 del. Cytological smears also showed EGFR mutations in the same pattern as tissue biopsies and a concordance was seen in cases where matched tissue and cytology smears were available.
Table1: EGFR mutation spectrum in pulmonary adenocarcinomas
EGFR Exon
Percentage
Single Mutations = 89.7% (210/234)
EGFR most common mutations
Exon 19 del
61.5%
Exon 21 L858R
29.1%
EGFR TKI resistant mutation at baseline
Exon 20 T790M
2.8%
Rare EGFR Single Mutations
Exon 18 G719X
2.4%
Exon 20 S768I
1.4%
Exon 21 L861Q
1.4%
Exon 20 Ins
1.4%
EGFR double mutations = 10.2% (24/234)
Common double mutations
Exon 19 del: Exon 20 T790M
11
Exon 19 del: Exon 21 L858R
5
Rare double mutations
Exon 18 G719X: exon 20 S768I
2
Exon 21 L858R: exon 20 S768I
2
Exon 20 T790M: Exon 18 G719X
1
Exon 20 T790M: Exon 20 Ins
1
Exon 19 del: Exon 21 L861Q
1
Exon 19 del: Exon 18 G719X
1
Conclusion
With judicious use and triaging of lung cancer diagnostic specimens, it is possible to perform successful mutation testing in >98% cases. EGFR mutation was positive in about a third of ADC including rare, TKI resistant and coexisting mutations in approximately 10% cases which may have significant therapeutic implications. In patients with limited amounts of tissue, cytology samples can be used for EGFR mutation testing as a promising alternative.
