Virtual Library
Start Your Search
M. Gallagher
Author of
-
+
P1.03 - Chemotherapy/Targeted Therapy (ID 689)
- Event: WCLC 2017
- Type: Poster Session with Presenters Present
- Track: Chemotherapy/Targeted Therapy
- Presentations: 2
- Moderators:
- Coordinates: 10/16/2017, 09:30 - 16:00, Exhibit Hall (Hall B + C)
-
+
P1.03-041 - Exploitation of the Cancer Stem Cell Marker ALDH1 Within the Vitamin a/Retinoic Acid Axis Promotes Re-Sensitisation of Cisplatin Resistant NSCLC (ID 9938)
09:30 - 09:30 | Author(s): M. Gallagher
- Abstract
Background:
Despite significant advances in personalised medicine in recent decades, cisplatin remains the mainstay chemotherapy in the treatment of NSCLC. The major clinical challenge facing NSCLC today is the development of pan-resistance to platinum agents. Novel drug design, preclinical and clinical trials working toward the approval of new drugs is a lengthy and costly process and in the interim of the drug indentifcation and commercialisation research has turned its focus to two avenues of interest; overcoming cisplatin resistance and the repurposing of approved therapeutics with new indications. Cancer stem cells (CSCs) have been hypothesised to be the initiating cells of therapeutic resistance and tumour recurrence. An ALDH1-positive cell subset has been identified as a key CSC subpopulation present within cisplatin resistant NSCLC sublines. ALDH1 is involved in the metabolism of retinol (vitamin A) and the catalytic conversion of retinal to retinoic acid, where retinoic acid induces cell differentiation. All-trans retinoic acid (ATRA) is a well-established chemotherapeutic agent in the treatment of acute promyelocytic leukaemia; it induces the terminal differentiation of immature cells. We hypothesise that treatment of the cisplatin resistant NSCLC sublines with retinol or ATRA will deplete the ALDH1-positive population and subsequently increase or restore cisplatin sensitivity.
Method:
Flow cytometry on a panel of matched parent (PT) and cisplatin resistant (CisR) NSCLC cell lines revealed the greater presence of ALDH1-positive CSC subpopulations within the CisR sublines of each NSCLC histology relative to PT lines. Cells were treated with retinol (substrate of the retinoic acid pathway) or ATRA (product of the retinoic acid pathway) and the presence of the ALDH1-positive CSC subset reanalysed by flow cytometry. Following treatment of the PT and CisR cells with retinol or ATRA alone and in combination with cisplatin the functional parameters of proliferation, clonogenic survival and apoptosis were reassessed relative to cisplatin alone.
Result:
Treatment of the CisR sublines with retinol (1μM) or ATRA (5μM) significantly reduced the presence of the ALDH1-positive CSC subset across CisR sublines. Both retinol and ATRA when used in combination with cisplatin significantly reduced the proliferative and survival capacity of each CisR subline while significantly increasing apoptotic cell death compared to cisplatin alone.
Conclusion:
Exploitation of the vitamin A/retinoic acid pathway in combination with cisplatin re-sensitised resistant cells to the cytotoxic effects of cisplatin. These data suggest vitamin A supplementation or the addition of FDA-approved ATRA to the cisplatin-based chemotherapeutic regimen may be of clinical benefit in overcoming tumour recurrence and cisplatin resistance.
-
+
P1.03-048 - miR-34a and the Micromanagement of Cancer Stemness and Resistance in NSCLC. Does It Hold Therapeutic Benefit? (ID 9968)
09:30 - 09:30 | Author(s): M. Gallagher
- Abstract
Background:
The capacity of microRNAs to post-transcriptionally regulate a myriad of genes has extended their remit into the realm of stemness and furthermore cancer stemness regulation. Disruption of Dicer-1, a crucial component of microRNA biogenesis, has been shown to completely deplete the stem cell pool in early development, indicating a potential role for microRNAs in the maintenance of stem cells. Such logic has led microRNAs to be investigated in the context of cancer stem cells (CSCs). Studies have revealed that microRNAs play a role in CSC self-renewal, differentiation, drug resistance and metastasis. With this, our hypothesis suggests that microRNAs associated with cisplatin resistance and CSC maintenance may be a key target by which the CSC root of cisplatin resistance could be overcome.
Method:
MicroRNA expression within a panel of age-matched parent (PT) and cisplatin resistant (CisR) NSCLC sublines was profiled using the 7[th] generation miRCURY LNA arrays (Exiqon) and validated by qPCR. Cell lines were stained for the presence of the CSC marker, aldehyde dehydrogenase 1 (ALDH1) and FACS was used to isolate the ALDH1-positive CSC population from the ALDH1-negative bulk cell population. Expression of the panel of cisplatin resistance-associated microRNAs was investigated within the ALDH1-positive CSC population relative to their negative counterparts by qPCR. Significantly altered miRNAs were inhibited in the CisR subline using antagomirs (Exiqon) and the presence of the ALDH1-positive subset reassessed by flow cytometry and expression of stemness genes (Nanog, Oct-4, Sox-2, Klf4, cMyc) determined. The presence of the cisplatin-associated miRNAs was investigated in FFPE murine tumours within a xenograft model of CSCs, in which 1x10[3] ALDH1-positive and negative subsets were injected into NOD/SCID mice.
Result:
Upon validation, a 5-miR signature was identified across NSCLC histologies to be associated with cisplatin resistance. When this panel was further investigated within the ALDH1-positive CSC subpopulation, it was observed that there was a significant up-regulation of miR-34a-5p relative to corresponding ALDH1-negative populations. Interestingly, the ALDH1-positive subpopulations showed significantly greater miR-34a-5p expression when compared to the CisR sublines from which they were isolated. This up-regulation was also observed within the FFPE xenograft tumours. However, inhibition of miR-34a-5p with antagomiRs did not significantly alter the presence of the ALDH1-positive CSC population, or the expression of stemness-associated genes.
Conclusion:
These data suggest that miR-34a-5p while significantly up-regulated in cisplatin resistance and CSCs may not play a functional role in CSC maintenance and further investigation is required to fully elucidate the role of miR-34a-5p in cancer stemness.
-
+
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)
-
+
P2.02-064 - A Novel 5-miR Signature Shows Potential as a Diagnostic Tool and as a Predictive Biomarker of Cisplatin Response in NSCLC (ID 9957)
09:30 - 09:30 | Author(s): M. Gallagher
- Abstract
Background:
MicroRNAs are a class of small non-coding RNAs that range in size from 19-25 nucleotides. They have been shown to regulate a number of processes within tumour biology, including metastasis, invasion and angiogenesis. More recently, miRNAs have been linked to chemoresistance in solid tumours, including lung cancer. Their role in cisplatin resistance has yet to be determined.
Method:
MicroRNA expression within a panel of age-matched parent (PT) and cisplatin resistant (CisR) NSCLC cell lines was profiled using the 7[th] generation miRCURY LNA arrays (Exiqon) and subsequently validated by qPCR. Significantly altered miRNAs within the CisR sublines were manipulated using antagomirs (Exiqon) and Pre-miRs (Ambion) and functional studies were carried out in the presence and absence of cisplatin. To examine the translational relevance of these miRNAs, their expression was examined in a cohort of chemo-naïve patient-matched normal and lung tumour tissue and serum from NSCLC patients of different histologies. A xenograft model of cisplatin resistance was carried out in which 1x10[3] H460 PT or CisR cells were injected into 5-7week old NOD/SCID mice. Tumour volume was measured over time and harvested once the tumour mass measured 500mm[3] and formalin-fixed and paraffin embedded (FFPE). Expression of the 5-miR signature was analysed within FFPE murine tumours and compared between PT and CisR tumours.
Result:
Profiling and subsequent validation revealed a 5-miR signature associated with our model of cisplatin resistance (miR-30a-3p, miR-30b-5p, miR-30c-5p, miR-34a-5p, miR-4286). Inhibition of the miR-30 family and miR-34a-5p reduced clonogenic survival of CisR cells when treated cisplatin. Expression of the miRNA signature was significantly altered in both adenocarcinoma (AD) and squamous cell carcinoma (SCC) relative to matched normal lung tissue and between SCC and AD tissue. miR-4286 was significantly up-regulated in SCC sera compared to normal control and AD sera. Similarly to the cell line expression of the miRNAs, the miR-30 family members and miR-34a-5p were up-regulated in the CisR xenograft FFPE tissue relative to PT.
Conclusion:
A novel miRNA signature associated with cisplatin resistance was identified in vitro, genetic manipulation of which altered clonogenic response to cisplatin. The 5-miR signature showed both diagnostic and prognostic biomarker potential across a number of diagnostically relevant biological media.
