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J. Jang
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P1.02 - Biology/Pathology (ID 614)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Biology/Pathology
- Presentations: 2
- Moderators:
- Coordinates: 10/16/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P1.02-015 - Comparison of Study Models of Lung Cancer (ID 9318)
09:30 - 09:30 | Author(s): J. Jang
- Abstract
Background:
Lung cancer is the most prominent cancer in human with high mortality rate. Although chemo-radiation therapies have been improved, the patient survival is still poor. Thus, not only developing therapeutic medications, but also biomarkers of early diagnosis have been desired. Several models of primary lung cancer research are in use, however, systematic evaluation and characterization of models are required to have suitability and relevance based on study aims. To provide research environment for lung cancer, we reappraise relevant models for lung cancer.
Method:
Three concepts of primary lung cancer models were evaluated: (I) Urethane induced lung tumor. (II) Cell line induced tumor model via intravenous (iv) or subcutaneous (sc) injection. (III) ex vivo 3D primary cell culture model. 20 weeks after urethane injection, the animals were harvested to analyze tumor incidence and tumor immunity. Lewis Lung Carcinoma (LLC) cell line was employed for the orthotopic development of lung tumor in 2 weeks after the injection. Hanging drop method was used for the 3D culture of primary cells from LLC sc induced tumor. Immunohistochemistry (IHC) of proliferation markers (pH3 and Ki67), tumor immunity (CD4, CD8, B220, F4/80, NKp46, and PDL-1) were performed for a finer characterization of tumors.
Result:
Urethane and iv injection of LLC cell line developed heterogeneously distributed tumors in lung. sc injection stably developed single tumor nodule. 3D cultured primary cells formed spheroids within 5 days. IHC revealed that all tumors were consistently proliferating with less extend in urethane and 3D model. F4/80[+ ]cells and CD4[+] cells infiltrated into tumors significantly more than CD8[+], B220[+], or NKp46[+] cells. T cell populations (CD4[+] and CD8[+]) were much more prominent in LLC iv model than other models. Interestingly the expression of PDL-1 was found only in 3D model.
Conclusion:
The urethane-induced primary lung tumor is reliable with a high rate of development, but needs longer time period to develop tumor compared to iv and sc models. iv injection model develops lung tumor in the original location. With relatively more convenience, sc model allows the analysis of tumor without adjacent tissue bias. 3D primary cell culture model enable for conferring characterization of individual tumor and strategic design of therapy, namely personalized medicine. The involvement and characteristics of immune cells found within tumors were comparable across all models. Injections by i.v. or s.c. of cell line to mouse can be considered as an alternative yet convenient model to develop various different types of lung cancers.
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P1.02-039 - Preventive and Therapeutic Action of Id1 Inhibition in KRAS-Mutant (KM) Lung Adenocarcinoma (LAC) Tumors in a Xenograft Murine Model (ID 9574)
09:30 - 09:30 | Author(s): J. Jang
- Abstract
Background:
Id1 has been shown to be involved in cell viability and migration of lung cancer cell lines and confer poor prognosis in LAC-patients. The most frequently mutation in LAC is KRAS, but no targeted therapy has been successfully developed. Here we study the role of Id1 in a KM-LAC murine model.
Method:
The expression of Id1 was analyzed in a panel of human LAC cell lines by qPCR and Western-Blot. Several human cell lines with known mutations (H1792-604, H2009, H358, H1568, H1437, H1703, H2126) were selected to deplete Id1 expression by inducible short hairpin RNA (shRNA) regulated by doxycycline. Proliferation, cell cycle and apoptosis assays were performed to study the cellular mechanism underlying the effect of Id1 deficiency. Mouse xenograft models were generated by subcutaneous injection of KM-LAC cells (H1792-604 and H2009), both shId1 and shGFP cells, in flanks of immunodeficient mice treated with doxycycline (drinking water) from the time of inoculation or once the tumors were established.
Result:
Id1 overexpression was observed in 11 out of 12 cell lines as occurs in previously reported clinical data. Id1 inhibition was achieved in all cell lines compared to controls. In absence of Id1, proliferation assays showed a significant impairment of cell growth in KM-LAC cell lines [H1792-604 31.61% ± 3.96 (P < 0.001); H2009 52.73% ±4.74 (P < 0.001); H358 70.85% ± 8.01 (P < 0.001)]. In KM cells, a significant arrest in G2/M phase of cell cycle was observed when Id1 was inhibited whereas no significant changes were observed in wild type(WT) KRAS cells [KM 1.86 ± 0.28;WT 1.02 ± 0.05 (P < 0.001)]. KM-cells showed a significant apoptosis increase compared to WT-cells [KM-cells 1.66 ± 0.41;WT-cells 0.99 ± 0.13 (P = 0.001)]. In vivo, we observed a significant decrease in tumor volume in mice injected with H1792-604-shId1 cells (60% ± 32.39) compared to shGFP group (356.29% ± 115.32)(P < 0.001). Moreover, mice injected with H2009-shId1 cells did not develop tumors compared to control mice (168.35 ± 68.71)(P < 0.001). Activation of shId1 in established tumors induced a significant reduction of tumor volume in both xenograft models. The inhibition led to regression of 4 out of 10 tumors H1792-604 and all tumors in H2009 inoculated mice.
Conclusion:
These findings support a crucial role of Id1 in tumor development in KRAS-driven adenocarcinoma of the lung. Id1 targeting was proven effective in both, tumor prevention and treatment in our humanized murine model of KM LAC.
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P2.02 - Biology/Pathology (ID 616)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Biology/Pathology
- Presentations: 1
- Moderators:
- Coordinates: 10/17/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
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P2.02-036 - The Expression Pattern of CD26/DPP4 in Human Lung Cancer (ID 9319)
09:30 - 09:30 | Author(s): J. Jang
- Abstract
Background:
Lung cancer is the leading cause of death among cancers. Despite improved surgical and novel radiation improvements, the overall prognosis remains poor. CD26/dipeptidyl peptidase 4 (DPP4) is a ubiquitously expressed transmembrane exopeptidase on the cell surfaces of many different cells including malignancies of breast, colon, and mesothelioma. Phase I data in mesothelioma with a specific antibody showed tolerability in mesothelioma patients.Our group found previously that the activity of CD26/DPP4 of lung adenocarcinoma (Adeno-CA) patients is four times higher than in normal tissue and the inhibition of CD26/DPP4 decreased the growth of lung tumors in experimental models. These data prompted us to analyze the expression of CD26/DPP4 in samples from lung cancer patients to unravel the role of CD26/DPP4 as a biomarker for lung cancer and a target for inhibition to reduce lung cancer burden. burden.
Method:
To identify CD26/DPP4 by immunohistochemistry (IHC), we tested four antibodies from Abcam, R/D systems, and Cell signaling technology on multi-organ tissue micro array (TMA) and human lung Adeno-CA cell lines (A549, H460, Gon8, Mai9) derived from advanced stage (IV) of human Adeno-CA. We selected the antibody from Cell signaling technology against CD26/DPP4. For the analysis of CD26/DPP4 by IHC in lung cancer samples, TMAs constructed from non-small cell lung cancer patients were used. The cohort consisted of 475 patients (Adeno-CA: 223; Squamous carcinoma: 252). The intensity of the staining was scored from 0 to 3 in a blinded manner. To quantify CD26/DPP4 in the supernatant of human lung Adeno-CA cell lines in vitro, ELISA was performed.
Result:
IHC scores revealed that Adeno-CA expresses significantly more CD26/DPP4 compared to squamous carcinoma (p<0.0001). Consistent with our previous findings, early stage cancer (IA) scores significantly higher than other stages IIB (p=0.0012), IIIA (p=0.0019), and IV (p=0.02) among Adeno-CA samples. We could not find CD26/DPP4 expression on human Adeno-CA cell lines by IHC, but the secretion of the protein in supernatant stays high (A549: 20pg/ml; H460: 161pg/ml; Gon8: 74pg/ml; Mai9: 648pg/ml).
Conclusion:
CD26/DPP4 expression was significantly higher at early stages of Adeno-CA samples when compared to advanced stages, supporting our previous findings. From the human cell line data, we suggest that advanced cancer secretes CD26/DPP4 more actively than early stage cancers. CD26/DPP4 seems to be a substantial target for inhibition of human Adeno-CA.
