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Don Lynn Gibbons



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    MA03 - Clinomics and Genomics (ID 119)

    • Event: WCLC 2019
    • Type: Mini Oral Session
    • Track: Advanced NSCLC
    • Presentations: 1
    • Now Available
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      MA03.05 - BRAF Mutations Are Associated with Increased Benefit from PD1/PDL1 Blockade Compared with Other Oncogenic Drivers in Non-Small Cell Lung Cancer (Now Available) (ID 1472)

      10:30 - 12:00  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Presentation
      • Slides

      Background

      PD-1/PD-L1 immune checkpoint blockade (ICB) has revolutionized the treatment of non-small cell lung cancer (NSCLC), but only a minority of patients achieve durable clinical benefit. Although classic EGFR/ALK alterations are correlated with ICB resistance, it is unknown if patients with other molecular subtypes of NSCLC also derive poorer outcomes from ICB. We investigated if there are oncogene-driven NSCLC associated with higher response rates (RR) and progression-free survival (PFS) to ICB.

      Method

      Two independent retrospective cohorts of oncogene-driven NSCLC treated with ICB monotherapy were analyzed for clinical outcome: MD Anderson (MDACC) and Flatiron Health-Foundation Medicine Clinico-Genomic Database (FH-CGDB). PD-L1 expression (Dako 22C3 - FoundationCore) and tumor mutational burden (TMB - FoundationCore; TCGA and MSK-IMPACT – cbioportal.org) were compared across distinct molecular subtypes of NSCLC to determine differences in clinical outcome.

      Result

      Among five oncogene defined groups from the MDACC cohort, BRAF-mutant NSCLC had the highest response rate (RR) (RECIST 1.1) (P<0.01) and PFS (P<0.01) when treated with ICB (Table). These differences remained significant after adjusting for PD-L1 expression. Classic EGFR and HER-2 mutant NSCLC had the lowest RR and PFS (Table). Similar results were observed in the independent FH-CGDB cohort where BRAF-mutant NSCLC had longer real-world (rw) PFS and OS to ICB monotherapy (Table). PD-L1 expression (tumor score ≥1% and ≥50%) and TMB were higher in BRAF-mutant NSCLC compared to EGFR and HER-2 (P<0.01). BRAF V600E NSCLC had lower TMB compared to non-V600E (5.9 vs 13.7 mut/Mb, P<0.01), but both had high PD-L1 expression (≥1%: 72% vs 61%; ≥50%: 42% vs 32%).

      KRAS

      BRAF

      Classic EGFR

      EGFR exon 20

      HER2

      MDACC cohort

      Patients – N

      87

      10 (V600E 3 / non-V600E 7)

      28

      25

      15

      RR – %

      24.3

      62.5

      4.5b

      10b

      8.3

      Median PFS – mo (95% CI)

      2.76

      (2.23-3.30)

      7.37 (not estimable)a

      1.78 (1.18-2.37)

      2.73 (1.71-3.75)

      1.88 (1.63-2.12)

      FH-CGDB

      Patients – N

      503

      68 (V600E 32 / non-V600E 36)

      52

      42

      25

      Median rwPFS -

      mo (95% CI)

      3.55

      (3.15-4.24)

      6.0

      (2.89-11.6)

      2.17b

      (1.77-2.63)

      2.66b

      (2.23-5.13)

      1.87b (1.31-4.34)

      Median rwOS – mo (95% CI)

      10.28

      (8.51-12.02)

      16.07

      (8.64-NA)

      5.29b

      (3.25-17.68)

      9.89b

      (3.68-20.86)

      10.81

      (4.17-NA)

      FoundationCore cohort – N

      NA

      188 (V600E 74 / non-V600E 114)

      386

      96

      57

      TMB – mean (mut/Mb)

      NA

      10.6a

      3.7

      3.8

      5.8

      PD-L1 TPS ≥ 50% (%)

      NA

      36a

      19

      23

      16

      a: P<0.01 vs all groups; b: P<0.05 for pairwise comparison vs BRAF.

      Conclusion

      NSCLCs with BRAF mutations are associated with increased benefit from ICB when compared to tumors harboring other targetable oncogenic drivers. Oncogene driver mutations are associated with distinct patterns of TMB and PD-L1 expression. These findings highlight the importance of developing mutation-specific clinical trials in NSCLC.

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    MA09 - EGFR & MET (ID 128)

    • Event: WCLC 2019
    • Type: Mini Oral Session
    • Track: Targeted Therapy
    • Presentations: 1
    • Now Available
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      MA09.03 - Identification of Mechanisms of Acquired Resistance to Poziotinib in EGFR Exon 20 Mutant Non-Small Cell Lung Cancer (NSCLC) (Now Available) (ID 2904)

      15:15 - 16:45  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Presentation
      • Slides

      Background

      Insertions/mutations in exon 20 of EGFR occur in ~2% Insertions/mutations in exon 20 of EGFR occur in ~2% of all lung adenocarcinomas. These alterations are characterized by primary resistance to approved tyrosine kinase inhibitors (TKIs) with response rates of <12%. We have shown that exon 20 insertions restrict the size of the drug-binding pocket, limiting binding of large inhibitors. However, poziotinib can circumvent these steric changes and is a potent inhibitor of EGFR exon 20 mutants. In our investigator-initiated phase 2 trial of EGFR exon 20 mutant NSCLC, poziotinib was associated with a best objective response rate of 55% (Heymach et al, 19th WCLC). Herein, we use preclinical models and clinical samples from our phase 2 study to identify mechanisms of acquired poziotinib resistance (NCT03066206).

      Method

      EGFR exon 20 insertion (D770insNPG) genetically engineered mice (GEM) were treated with poziotinib until progression. Upon progression, tumor DNA and protein were analyzed using whole exome sequencing (WES) and reverse phase protein assay (RPPA). Mandatory and optional biopsies were obtained at baseline and progression, respectively, from patients treated in our phase 2 trial of poziotinib in EGFR exon 20 mutant NSCLC. Serial cfDNA was collected at baseline, 8 weeks of therapy, and on progression. Patient samples were analyzed using targeted next generation sequencing or WES.

      Result

      Poziotinib acquired-resistance GEM tumors acquired mutations in ErbB4, KRAS, and other genes which represent potential targetable bypass pathways. Resistant GEM tumors displayed increased activation of MAPK, AKT, ERK and MEK compared to sensitive tumors, suggesting that poziotinib acquired resistance is associated with reactivation of the MAPK/PI3K pathways. We enrolled 50 EGFR exon 20 mutant patients in our phase 2 trial. Analysis of matched pre-poziotinib and on-progression samples from 20 responding patients revealed acquired EGFR tyrosine kinase domain point mutations in 4 patients (T790M (2), V774A (1), D770A, (1)). Ba/F3 cells co-expressing EGFR exon 20 insertion (S768supSVD) and T790M were resistant to poziotinib, suggesting that T790M is a poziotinib resistance driver. Potential acquired EGFR-independent resistance mechanisms identified in patients to date include PIK3CA E545K (1), MAP2K2 S94L (1), MET amplification (1), EGFR amplification (2), and CDK6 amplification (2).

      Conclusion

      Parallel to acquired resistance mechanisms seen in classical EGFR mutation, acquired resistance to poziotinib can be mediated through EGFR-dependent mechanisms, notably T790M and other EGFR tyrosine kinase domain point mutations. EGFR-independent resistance mechanisms include activation of bypass pathways. Preclinical validation of resistance mechanisms and additional analysis of patient samples will be presented at the meeting.

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    MA11 - Immunotherapy in Special Populations and Predictive Markers (ID 135)

    • Event: WCLC 2019
    • Type: Mini Oral Session
    • Track: Immuno-oncology
    • Presentations: 1
    • Now Available
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      MA11.09 - Increased Frequency of Bystander T Cells in the Lungs Is Associated with Recurrence in Localized Non-Small Cell Lung Cancer (Now Available) (ID 955)

      14:00 - 15:30  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Presentation
      • Slides

      Background

      Non-small cell lung cancer (NSCLC) exhibits a high mutational burden. As a result, patients afflicted by this tumor type experience greater responses to immune checkpoint blockade. This is largely due to the ability of T cells to destroy tumor cells on the basis of antigens recognized by their T cell receptor (TCR). However, the lungs are exposed to carcinogens and pathogens which can also trigger a T cell response distinct from cancer. Therefore, a better understanding of the T cell repertoire in the lungs is needed to improve upon the success of current immunotherapies in NSCLC.

      Method

      We obtained peripheral blood, tumors, and adjacent uninvolved lungs from a cohort of 236 early stage NSCLC patients. Whole exome sequencing, RNA microarray, immunohistochemistry (CD3, CD4, CD8, CD57, CD68, FoxP3, CD45RO, GzmB, PD-1, and PD-L1) and T cell repertoire sequencing were performed in NSCLC patients and lungs from organ donors and COPD patients. Antigen specificity was predicted using the Grouping of Lymphocyte Interactions by Paratope Hotspot (GLIPH) algorithm. Single cell TCR and RNA sequencing as well as sequencing of the virome are underway.

      Result

      Clonality was associated with CD8 T cells (r=0.31; p=0.0003), GzmB (r=0.29; p=0.001) and IFN-γ (r=0.52; p<0.0001) production as well as with tumor mutational burden (r=0.19; p=0.015), HLA-B (r=0.29; p=0.0005) and β2-m expression (r=0.20; p=0.018). Patients with classical EGFR mutations exhibited lower T cell clonality (p=0.003) even after adjustment for TMB, highlighting the impact of this driver mutation on the T cell response. Surprisingly, clonality was higher in the adjacent uninvolved lung than tumor (p<0.0001), suggesting an active antigenic response outside the tumor. Comparison of the composition of the T cell repertoire between the uninvolved lung and tumor revealed 57% of the top 100 T cells in the tumor were also found in the adjacent normal lung, highlighting certain parallels in the ongoing antigenic responses. Deeper analysis suggested that shared T cells may have been reactive against mutations shared between the normal lung and tumor (r=0.23, p=0.028) or viruses (p<0.0001). Accordingly, patients with a more reactive T cell repertoire outside the tumor (i.e. bystanders) exhibited shorter disease-free survival (p=0.036) suggesting these responses against shared mutations and/or viruses may detract from the anti-tumor T cell response.

      Conclusion

      Our findings highlight the importance of understanding the specificity of the T cell repertoire in the lungs in patients with NSCLC treated with immunotherapy. As a high proportion of bystander T cells appear to reside in the lungs, their reactivation could contribute to the impaired responses and/or increased toxicity observed in certain patients with NSCLC treated with immunotherapy.

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    OA13 - Ideal Approach to Lung Resection and Novel Perioperative Therapy (ID 146)

    • Event: WCLC 2019
    • Type: Oral Session
    • Track: Treatment of Early Stage/Localized Disease
    • Presentations: 1
    • Now Available
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      OA13.06 - Surgical Outcomes Following Neoadjuvant Nivolumab or Nivolumab Plus Ipilimumab in Non-Small Cell Lung Cancer - NEOSTAR Study (Now Available) (ID 2041)

      11:30 - 13:00  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Presentation
      • Slides

      Background

      Surgical outcomes following neoadjuvant immune checkpoint inhibitors (ICIs) are limited. We report 90-day perioperative results of the NEOSTAR phase II trial of neoadjuvant nivolumab or nivolumab/ipilimumab in resectable non-small cell lung cancers (NSCLCs).

      Method

      44 pts with stage I-IIIA NSCLC (AJCC 7th) were randomized to nivolumab (3 mg/kg IV, days 1, 15, 29, n=23) or nivolumab/ipilimumab (1 mg/kg IV, day 1, n=21) with resection planned between 3-6 weeks after last dose. Surgical approach and extent of resection were at surgeons’ discretion.

      Result

      39 (89%) patients underwent R0 resection, of those 2 (5%) were resected off trial after additional induction chemotherapy (1 nivolumab, 1 nivolumab/ipilimumab). Among 37 patients, 21 underwent surgery following nivolumab and 16 following nivolumab/ipilimumab. Median age 66 (43-83) years, 24 (65%) male, 33 (89%) white, 22 (59%) adenocarcinoma, 22 (59%) stage I, 9 (24%) stage II, 6 (16%) stage IIIA.

      5 (11%) were not resected, 1 (1/23, 4%) after nivolumab (stage II), 4 (4/21, 19%) after nivolumab/ipilimumab (1 stage I, 1 stage II, 2 stage IIIA). Reasons for unresectability were change in surgeon’s judgement (n=2), toxicity (n=1), progression (n=1), and declining pneumonectomy (n=1). Median time to surgery was 31 days (range 21-87). 8 (22%) operations were delayed beyond 42 days, 5 after nivolumab/ipilimumab (5/16, 31%) and 3 after nivolumab (3/21, 14%).

      33 (89%) underwent lobectomy, 2 (5%) pneumonectomy, 1 (3%) segmentectomy and 1 (3%) wedge resection. 27 (73%) had thoracotomy, 7 (19%) thoracoscopy, 3 (8%) robotic approach. 2 (5%) were electively converted from thoracoscopy to thoracotomy. Median operative time was 147 minutes (71-315), median blood loss was 100cc (50-1000), and median length of stay was 4 days (1-18).

      Perioperatively, pulmonary complications occurred in 8 (22%) patients: 8 (22%) prolonged air leak, 2 (5%) pneumonitis/pneumonias, 1 (3%) empyema, and 1 (3%) bronchopleural fistula (BPF). 1 (3%) died from complications of BPF and steroid therapy for pneumonitis. 4 (11%) developed atrial fibrillation, 1 (3%) diarrhea, 1 (3%) ileus, and 1 (3%) transient ischemic attack.

      Surgeons subjectively judged 15/37 (40%) of operations to be more complex than usual with 7/37 (19%) lasting > 4 hours.

      Conclusion

      Following three cycles of neoadjuvant ICIs 89% of patients underwent complete R0 resection, including two patients who received additional induction chemotherapy off trial. Five marginally operable patients who didn’t proceed to resection, and one perioperative mortality highlight the importance of cautious patient selection for neoadjuvant ICIs in the management of operable NSCLC.

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    OA15 - Targeted Agents and Immunotherapy for Small Cell Lung Cancer (ID 152)

    • Event: WCLC 2019
    • Type: Oral Session
    • Track: Small Cell Lung Cancer/NET
    • Presentations: 1
    • Now Available
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      OA15.04 - Genomic and TCR Intratumor Heterogeneity of Small-Cell Lung Cancer by Multiregion Sequencing: An Association with Survival (Now Available) (ID 1458)

      14:30 - 16:00  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Presentation
      • Slides

      Background

      Small cell lung cancer (SCLC) is an aggressive cancer. Although sensitive to initial therapy, recurrence is almost inevitable. The molecular mechanisms underlying recurrence are unknown. We have previously demonstrated that complex genomic and T cell receptor (TCR) intratumor heterogeneity (ITH) was associated with increased risks of relapse in non-small cell lung cancers (NSCLC). Genomic ITH and TCR architecture of SCLC and its clinical impact have not been well studied, largely due to lack of tumor specimens as surgery is rarely used to treat SCLC.

      Method

      We performed multiregion whole-exome sequencing and TCR sequencing of 49 tumor samples from 18 resected limited-stage SCLCs to delineate the immunogenomic ITH of SCLC. We compared the results to those in NSCLC and assessed the association of genomic and TCR attributes with patient’s survival.

      Result

      On average, 544 mutations/sample were detected. The median proportion of trunk mutations (mutations identified in all regions within the same tumors) was 80.4% versus 70% in NSCLC (TRACERx, Jamal-Hanjani, NEJM, 2017, p=0.08) and all TP53 and RB1 mutations were trunk mutations, suggesting these mutations were early events during carcinogenesis of this cohort of SCLCs. A higher non-synonymous tumor mutational burden (TMB) was associated with a higher T cell density (infiltration) in the tumor (r=0.46, p=0.005). Compared to the TCR repertoire of NSCLC (Reuben, WCLC, 2017), these SCLC tumors demonstrated significantly lower T-cell density (0.05 versus 0.24, p<0.0001), richness (diversity, 1,043 versus 3,666, p<0.0001) and clonality (reactivity, average 0.02 versus 0.15, p<0.0001) despite similar non-synonymous TMB (average 187 in SCLC versus 176 mutations/sample in NSCLC). Only 0.2% to 14.6% of T cells were detectable across all regions from the same tumors, suggesting substantial TCR ITH. Jaccard index (JI), a parameter quantifying TCR ITH was significantly lower in SCLC than in NSCLC (0.06 versus 0.1, p<0.0001) implying higher level of TCR ITH in SCLC than NSCLC. Interestingly, higher T-cell density, richness or clonality appeared to be associated with lower risk of recurrence numerically. Furthermore, higher TCR JI (less degree of ITH) was associated with significantly longer overall survival (HR=0.15, p=0.04).

      Conclusion

      Limited-stage SCLC tumors have distinct TCR repertoire and genomic ITH architecture. Overall, SCLC may have a more pronounced immunosuppressive microenvironment and higher level of TCR repertoire ITH than NSCLC. Nevertheless, higher degree of T cell infiltration and clonal expansion as well as more homogeneous T cell response may be associated with more favorable clinical outcome in patients with limited-stage SCLC.

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    P1.01 - Advanced NSCLC (ID 158)

    • Event: WCLC 2019
    • Type: Poster Viewing in the Exhibit Hall
    • Track: Advanced NSCLC
    • Presentations: 1
    • Moderators:
    • Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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      P1.01-98 - Outcomes in Advanced NSCLC Patients Treated with 1st Line EGFR-TKI Based on Mutation Detection from Tissue or cfDNA-Based Genomic Sequencing (ID 1861)

      09:45 - 18:00  |  Author(s): Don Lynn Gibbons

      • Abstract

      Background

      Tumor genomic information from tissue has been the standard of practice for identifying actionable molecular alterations. The same genomic profiling is also widely available by a non-invasive blood test (cfDNA). We hypothesized that treatment naïve patients with advanced non-small cell lung cancer (NSCLC) and actionable oncogenic driver mutations identified by tumor and cfDNA would have similar clinical outcomes after treatment with targeted therapies.

      Method

      Patients with any EGFR-TKI sensitive mutation and received FDA-approved EGFR-TKI as first line therapy for their advanced NSCLC were included in this retrospective analysis. Consecutive patients were identified from our GEMINI database with therapy initiated that was based solely from either the tissue or cfDNA report were divided into each cohort, respectively. Assessment of PFS was from date of therapy initiation until disease progression. Tissue genomic profiling was performed on our institution’s CLIA-certified hotspot NGS assay covering 40-50 genes. For blood based genomic profiling, blood was sent for NGS of cfDNA with a panel of up to 70 cancer-related genes at a CLIA-certified lab (Guardant360, Guardant Health, Redwood City, CA). Kaplan–Meier methodology was used to calculate median PFS with Log-rank (Mantel-Cox) test assessment at significance level 5%.

      Result

      Forty patients for each group were identified between 2014-2016. The results as summarized in table and PFS graph below:

      table.jpgpfs graph.jpg

      Conclusion

      There was no progression-free survival difference in patients treated with FDA-approved front-line EGFR-TKI directed by genomic profiling from tissue vs blood -based testing. These results indicate that similar treatment outcomes with targeted therapy based on tissue or blood-based NGS profiling are both viable options for patient with newly diagnosed, advanced NSCLC.

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    P1.04 - Immuno-oncology (ID 164)

    • Event: WCLC 2019
    • Type: Poster Viewing in the Exhibit Hall
    • Track: Immuno-oncology
    • Presentations: 4
    • Moderators:
    • Coordinates: 9/08/2019, 09:45 - 18:00, Exhibit Hall
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      P1.04-07 - Immune Suppressive Microenvironment and Highly Clonal Concordance of TCR Repertoire in Brain Metastases from Non-Small Cell Lung Cancer (ID 2018)

      09:45 - 18:00  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Slides

      Background

      The tumor immune microenvironment (TIME) of lung cancer brain metastasis is largely unexplored. We performed immune profiling and sequencing analysis of paired resected primary tumors and brain metastases of non-small cell lung carcinoma (NSCLC).

      Method

      TIME profiling of archival formalin-fixed and paraffin embedded specimens of paired primary tumors and brain metastasis from 39 patients with surgically resected NSCLCs was performed using a 770 immune gene expression panel (NanoString Technologies, Seattle, WA) and by T cell receptor beta repertoire (TCRß) sequencing (Adaptive Biotechnologies, Seattle, WA). Immunohistochemistry was performed for validation. Targeted sequencing was performed to catalog hot spot mutations in cancer genes (ThermoFisher Scientific, Waltham, MA).

      Result

      Somatic hot spot mutations were mostly shared between both tumor sites (28/39 patients; 71%). We identified 161 differentially expressed genes, indicating inhibition of dendritic cell maturation, Th1, and leukocyte extravasation signaling pathways, in brain metastases compared to primary tumors (p < 0.01). The proinflammatory cell adhesion molecule vascular cell adhesion protein 1 was significantly suppressed in brain metastases compared to primary tumors. Brain metastases exhibited lower T cell and elevated macrophage infiltration compared with primary tumors (p < 0.001). T cell clones were expanded in 64% of brain metastases compared with their corresponding primary tumors. Further, while TCR repertoires were largely shared between paired brain metastases and primary tumors, T cell densities were sparse in the metastases.

      Conclusion

      We present findings that the TIME in brain metastases is immunosuppressed when compared to matched primary tumors in NSCLC patients, and that thus may help guide immunotherapeutic strategies for NSCLC brain metastases.

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      P1.04-11 - Depicting the Intra-Tumoral Viral and Microbial Landscape of Localized NSCLC Using Standard Next Generation Sequencing Data (ID 1126)

      09:45 - 18:00  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Slides

      Background

      Studies from our group and others have shown that bacteria and viruses present in the tumor may impact therapeutic responses. In the specific context of non-small cell lung cancer (NSCLC), intra-tumoral viral DNA and bacteria have been reported previously to be linked to therapeutic outcomes. However, the interplay between intra-tumoral microorganisms and the host immune response in NSCLC remains unknown. Moreover, the prognostic and predictive therapeutic value of localized NSCLC-specific microbial composition has yet to be defined.

      Method

      RNA-sequencing (RNA-seq) (n=82) and whole exome sequencing (WES) (n=80) was performed on surgically resected (pTNM I-III) tumors from lung cancer patients enrolled in the ImmunogenomiC prOfiling of NSCLC (ICON) project. Intra-tumoral bacteria, viruses and fungi were queried with MetaPhlAn2, a bioinformatical analysis pipeline which employs unique clade-specific marker genes, using reads from RNA-seq and WES that did not map to the human genome/transcriptome. Generated data were correlated to patients’ clinicopathologic parameters as well as immune profiling using previously validated multiplex IHC panels based on Vectra 3.0™ multispectral microscopy IHC panels and image analysis (InForm™ 2.2.1 software).

      Result

      Our analyses revealed that 18.29% (n=15/82) of tumors contained bacterial signatures. The most frequent bacterial signature was related to Escherichia (n=9/15). Moreover, 6.49% (n= 5/77) of tumors had evidence of human viral signatures, including the Epstein-Barr virus (n=1/5). No tumors contained fungal signatures. Preliminary clinicopathologic analyses suggested that patients whose tumors harbor bacterial signatures had a trend towards decreased overall survival (p=0.12). Tumors from former smokers were also more likely to contain bacterial signatures (p=0.11). Preliminary multiplex immune cell IHC analyses did not highlight statistically significant associations with the presence of intra-tumoral bacteria.

      Conclusion

      Our results suggest that a significant proportion of localized NSCLC tumors may harbor components of the human microbiome. Further studies using larger cohorts and dedicated intra-tumoral microbiome and virome methodologies will be needed to better define these findings and to delineate associations with the local immune infiltrate.

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      P1.04-26 - EMT-Associated Response and Resistance to MEK Inhibitor and Immune Checkpoint Blockade Combinations in KRAS-Mutant NSCLC (ID 1129)

      09:45 - 18:00  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Slides

      Background

      Current work by our group using mutant KRAS and TP53 (KP) mouse models of NSCLC have shown that rationally designed therapies combining PD-L1 immune checkpoint blockade (ICB) with MEK inhibitors (MEKi) significantly decreases tumor growth and metastases compared to either monotherapies in syngeneic KP mice tumors. Despite these encouraging results, therapeutic resistance still occurs. Analyses from these tumors showed an increase in Tregs and CTLA-4 immune checkpoint expression. As anti-CTLA-4 ICB is particularly effective in increasing the CD8 / Treg ratio, we hypothesized that the addition of this agent may improve the outcome.

      Method

      Using in vivo KP syngeneic mouse models, we compared tumor size, tumor weight and lung metastatic nodules between two treatment regimens: the triple combination of selumetinib (MEKi) and anti-PD-L1 with either: 1) anti-CTLA-4 or; 2) IgG2b isotype control. FACS-based immunoprofiling was conducted at the time of response (5 weeks following treatment initiation) and resistance (maximal tumor volume). Whole tumors at the time of response and resistance, as well as ex vivo resistant cell lines, were also characterized by qPCR and Western Blotting (WB). Moreover, whole tumors from multiple treatment combinations and KP models were processed for custom codeset Nanostring mRNA analyses.

      Result

      The addition of anti-CTLA-4 to anti-PD-L1 and MEKi improved survival in the epithelial 393P KP mouse model (HR=3.517; p=0.03). Because FACS immunoprofiling of cytotoxic CD8+ T cells subtypes, NK cells and Tregs did not reveal statistically significant changes (p>0.05), we investigated potential tumor-intrinsic mechanisms. All resistant 393P cell lines displayed a mesenchymal morphology. Furthermore, whole tumors from the anti-CTLA-4 group demonstrated significantly less expression of Zeb1 (WB; p=0.05) at the time of response. Nanostring analyses comparing anti-PD-1 + anti-CTLA-4 vs anti-PD-L1 monotherapy in 344SQ KP mesenchymal tumors also showed statistically significant downregulation of epithelial-to-mesenchymal (EMT) markers (p=0.001). Finally, in vivo experiments using resistant 393P (MEKi + ICB) and mesenchymal 344SQ cells, demonstrated abrogation of the survival benefit initially observed with sensitive epithelial 393P cells upon treatment with combination therapies.

      Conclusion

      The combination of a MEKi, anti-PD-L1 and anti-CTLA-4 improves survival in epithelial syngeneic KP pre-clinical models of NSCLC, and this benefit is associated with downregulation of EMT markers. Therefore, further in-depth studies are required to understand the effect of ICB on EMT. In an upcoming single center, Phase I / II clinical trial, two combination schedules of selumetinib, tremelimumab and durvalumab will be compared with historical controls in patients with previously treated, metastatic NSCLC.

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      P1.04-79 - CD73 Expression in Lung Adenocarcinomas and Immunological and Molecular Associations (ID 2412)

      09:45 - 18:00  |  Author(s): Don Lynn Gibbons

      • Abstract

      Background

      Immune checkpoints inhibitors (ICI), in monotherapy or combination with chemotherapy, are the standard of care for lung adenocarcinoma (ADC) patients. Unfortunately, only a restricted number of patients will respond to ICI. Combination therapies such as CD73 inhibitors, are being studied with the goal to achieve synergic effects. CD73 is a membrane-bound protein with immunosuppressive functions. We previously reported that higher immune cell infiltration was associated mainly to CD73 basolateral (BL) expression, in this abstract, we show the correlation of CD73 expression at luminal (L) and BL membrane of ADC malignant cells (MCs), with annotated clinicopathological characteristics, immune and molecular biomarkers.

      Method

      CD73 IHC expression (clone D7F9A) was evaluated in 106 archived ADCs from patients that underwent surgical treatment without neoadjuvant therapy between February 1999 and February 2012 at MD Anderson Cancer Center (Houston, Texas, USA). We scored % and H-score of CD73 expression at the luminal (L) and basolateral (BL) membrane, we calculated the Total (T) CD73 as the average of L and BL, and classified ADCs in three groups: ‘T High’ (TH) (upper quartile for all tumors); ‘T Low’ (TL); ‘T Neg’ (TN) (<1%). We correlated T, L and BL expression and the three groups with clinicopathological characteristics, mutational status of KRAS and EGFR, TP53, STK11 and Tumor mutation burden (TMB), and cell densities of CD3, CD8, CD68, CD45RO, FOXP3, and Granzyme B, and PD-L1 expression (clone E1L3N) in MCs.

      Result

      T CD73 expression was found in 76%; BL in 60% and L in 57%; among ADCs with luminal membrane present (n=72), L CD73 was present in 83%. T+ and L+ expression was more frequent in never smokers (p=0.02 and p=0.003). Also higher frequency of L+ was found in older patients (>65) (p=0.01), tumors with non-solid histology patterns (p<0.001), EGFR mutation (p=0.048), non-mutated p53 (p=0.002), negative PD-L1 (p=0.03), and low TMB (<10 mut/MB) (p=0.001). Higher levels of L expression were found in KRAS mutated tumors (p=0.049). Higher BL expression positively correlated with p53 mutated tumors (p=0.038), PD-L1+ in MCs (p=<0.0001), and higher TMB (p=0.040).

      Our group analyses revealed that TH and TN were associated with ADCs from patients with >30 pack-year of smoking history (p=0.04), presence of any-solid histology pattern (p=0.03), p53 mutation (p= 0.005) and higher TMB (p=0.003) compared with TL. TH also had higher frequency of PD-L1+ tumors, and a higher cell density of CD3 (p=0.0001), CD8 (p=0.001), CD68 (p=0.048), CD45RO (p=0.036), FOXP3 (p=0.053), and Granzyme B (p=0.024) compared to TL and TN. TN showed higher frequency of STK11 mutation (p=0.034).

      Conclusion

      Based on the CD73 expression we defined subsets of lung adenocarcinomas that have distinct histological, molecular and immunological characteristics that may play a role in the response to ICI.

      Our characterization could help us to understand patient’s response to ICI, and identify patients that could potentially benefit from combination therapies.

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    P2.04 - Immuno-oncology (ID 167)

    • Event: WCLC 2019
    • Type: Poster Viewing in the Exhibit Hall
    • Track: Immuno-oncology
    • Presentations: 2
    • Moderators:
    • Coordinates: 9/09/2019, 10:15 - 18:15, Exhibit Hall
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      P2.04-19 - Neoadjuvant Chemotherapy Is Associated with Immunogenic Cell Death and Increased T Cell Infiltration in Early-Stage NSCLC (ID 1122)

      10:15 - 18:15  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Slides

      Background

      Recent success using immune checkpoint blockade (ICB) in the metastatic setting has raised the need to understand the immune microenvironment (IME) in early-stage disease. Moreover, pre-clinical evidence suggests that cytotoxic agents can modulate this IME. A recent study conducted by our group showed that non-small cell lung cancer (NSCLC) patients who received neoadjuvant chemotherapy followed by surgery (NCT), as compared to patients who received upfront surgery (US), had higher densities of CD3+ lymphocytes and CD68+ tumor-associated macrophages (TAMs). CD3+CD4+ lymphocytes and TAMs also correlated with better clinical outcomes. In this study, we explored the relationships between NCT and the IME by harvesting tumor samples of multiple surgical NSCLC cohorts.

      Method

      The PROSPECT microarray database was queried in NCT (n=45) and US (n=200) patients to investigate differentially expressed genes related to immunogenic cell death (ICD), susceptibility to CD8+ T cell and NK cell cytotoxicity, priming of antigen presenting cells, immunosuppressive enzymes and intra-tumoral cytokines. Available data from the ImmunogenomiC prOfiling of NSCLC (ICON) and other surgical NSCLC cohorts was evaluated to determine: 1) differential immune profiling using FACS (NCT=17; US=39) and multiplex IHC imaging (NCT=10; US=72); 2) plasma circulating cytokines (NCT=18; US=73); 3) tumor mutational burden (TMB) (NCT=40; US=61). Participants who received NCT or US were excluded according to these criteria: 1) concurrent treatment in addition to NCT; 2) sarcomatoid and small cell histologies; 3) clinical or pathological TNM Stage 4 disease; 4) synchronous malignancies other than lung.

      Result

      PROSPECT NCT patients expressed increased damage-associated molecular pattern (DAMP) genes (HSPA2, HSPA4, HSPE1, and S100A2; p<0.05) and T cell-related chemotaxis and antigen presentation genes (CXCR7, CD1A; p<0.05). Concordantly, the ICON cohort FACS results showed that NCT patients display increases in: 1) infiltration of CD8+ T cells (p=0.004); 2) proliferating Ki67+CD8+ T cells (p=0.02); 3) tissue resident memory CD8+CD103+ (p=0.02) and CD4+CD103+ non-Treg cells (p=0.01). Trends from the ICON multiplex IHC also highlighted increases in CD8+ T cells (p=0.09), CD20+ cells (p=0.08), as well as PD-L1+ malignant cells (p=0.08) and PD-L1+ TAMs (p=0.08) in NCT patients, the latter finding being supported by increased circulating MCP-1 (p=0.03). TMB was similar between NCT and US groups (p=0.912).

      Conclusion

      Our data provides the first evidence of ICD (i.e., increased DAMP gene expression) following NCT in human early-stage NSCLC. Furthermore, our data highlights the association of NCT with a favorable IME (i.e., increased T cell infiltration), supporting the rationale of NCT and ICB combinations in localized NSCLC.

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      P2.04-37 - Phase I/II Trial of Durvalumab and Tremelimumab with Continuous or Intermittent MEK Inhibitor Selumetinib in Advanced NSCLC (ID 1130)

      10:15 - 18:15  |  Author(s): Don Lynn Gibbons

      • Abstract
      • Slides

      Background

      Despite therapeutic progress in other molecular subsets of non-small cell lung cancer (NSCLC), little progress has been made for KRAS-mutant NSCLC. Because RAS remains an elusive pharmacological target, agents targeting downstream elements of the MAPK signaling pathway have been developed, including MEK inhibitors. However, substantial benefits have not been achieved due to the development of drug resistance. Current strategies to improve outcomes in this population include MEK inhibitors and PD-1 / PD-L1 immune checkpoint blockade (ICB) combinations (e.g., NCT03225664). On the other hand, combined anti-CTLA-4 and PD-1 / PD-L1 axis ICB improves response rates in melanoma, but similar benefits remain to be seen in NSCLC: MYSTIC trial updates failed to show a progression-free survival (PFS) advantage over standard of care, and CheckMate 227 reported significantly longer PFS with first-line double ICB in the high tumor mutational burden subgroup only. Thus, the objectives of this study are: 1) to determine the safety and efficacy of combined MEK inhibition, anti-PD-L1 and anti-CTLA-4 and; 2) to unveil mechanistic insights for response and resistance.

      Method

      This is a single center, Phase I/II study comparing two combination schedules of selumetinib (AZD6244, ARRY-142886), tremelimumab and durvalumab with historical controls in patients with previously treated, unresectable NSCLC. Forty patients will be accrued at the University of Texas MD Anderson Cancer Center. In the first arm, participants receive selumetinib PO BID on days 1-7 and 15-21 and durvalumab IV on day 1. Participants also receive tremelimumab IV on day 1 for courses 1-4 (courses repeat every 28 days in the absence of disease progression or unacceptable toxicity). The second arm differs by the continuous selumetinib schedule: PO BID on days 1-28. Primary objectives include the maximum tolerated dose (MTD; dose-escalation phase) and PFS (dose expansion phase). Standard 3+3 design will be applied to determine the MTD among the three pre-defined dose levels. Estimated PFS will be provided with 95% confidence interval. Secondary objectives include: 1) response rate by RECIST 1.1; 2) disease control rate (complete response + partial response + stable disease); 3) overall survival; 4) safety and; 5) duration of response. Exploratory objectives will assess biomarkers of response and resistance in pre- and on-treatment biopsies as well as peripheral blood using immune profiling, transcriptome and protein readouts.

      Result

      Section not applicable (Clinical Trial in Progress)

      Conclusion

      The estimated start date for this trial (NCT03581487) is April 15th, 2019, and the estimated study completion is scheduled for April 2021.

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