Page 152 - 2014 Printable Abstract Book
P. 152
(PS2-28) Potential role of miR-125a-5p as an indicator of radio response and treatment outcome in high
1
risk head and neck cancers after post-operative radio therapy. Narasimha Kumar Karanam, PhD ; Liang-
2
1
1
2
Hao Ding, PhD ; Uma Giri, PhD ; John V. Heymach, MD PhD ; and Michael D. Story, PhD, UT Southwestern
Medical Center, Dallas, TX and MD Anderson Cancer Center, Houston, TX
2
1
The use of miRNA as a biomarker has been proposed in a number of biological and medical
conditions such as growth and development, cancer development and metastasis, the cellular stress
response, and the radioresponse of cells and tissues. The advantages of miRNA as a biomarker lies in its
stability in tissues as well as body fluids, hence the potential for non-invasive diagnosis and prognosis. In
this study, we performed miRNA expression profiling using tumor specimens from 112 head and neck
cancer patients considered at high risk for recurrence after treatment with post-operative radiotherapy.
Specimens were triaged based upon patient outcome: no evidence of disease (NED), local recurrence (LR),
and distant metastasis (DM). Total RNA was extracted using a modified protocol of the Qiagen RNeasy
Plus kit which retains small RNA fractions. Labeled cRNAs were hybridized using the Exiqon whole genome
miRNA chip V17. Comparisons of miRNA expression between DM and NED specimens using a two-sample
t test identified 94 miRNAs were differentially expressed with statistical significance (p < 0.05), whereas
45 miRNAs were differentially expressed with statistical significance (p < 0.05) between LR and NED
specimens. Our hypothesis is; differentially expressed miRNA in the LR group would be associated with
clinical radioresistance and could serve as therapeutic targets. Amongst the miRNAs that were
significantly differentially expressed between LR and NED, we focused on miR-125a-5p as it had been
identified as a negative prognostic indicator in gastric cancers, and it targeted the ERBB2 gene. Down-
regulation of ERBB2 has been shown to result in inhibition of tumor growth and radiosensitivity in lung
and colorectal tumor cells. MiR-125a-5p expression was confirmed by qRTPCR, that is, down-regulated in
LR specimens as compared to that seen in NED specimens. Transfection of miR-125a-5p mimics into HN5
head and neck cancer cells inhibited cell proliferation and increased radiosensitivity about 30% after a 2
Gy dose, whereas miR-125-5p inhibitors increased cell proliferation and increased radio resistance about
90% after a 2 Gy dose. We are expanding our findings in a larger series of head and neck cancer cells and
confirming the regulation of ERBB2 by miR-125a-5p.
(PS2-29) The role of ATM, Apollo and MRE11 in the sensitivity of telomeric regions to DNA double-
strand breaks. Keiko Muraki; Douglas Miller; John P. Murnane, Univ. of Calif., San Francisco, San Francisco,
CA
Telomeres can interfere with DNA repair, as shown by an increase in large deletions and gross
chromosome rearrangements (GCRs), and a decrease in NHEJ at telomeric DSBs. In normal somatic cells,
the sensitivity of telomeric regions to DSBs is important in ionizing radiation-induced cell senescence and
in protection against oncogene-induced replication stress. However, in cancer cells or embryonic stem
cells, we have shown that DSBs near telomeres result in genomic instability. We previously reported that
a deficiency in ATM further increases the already high frequency of large deletions at telomeric DSBs, and
further decreases the already low frequency of distal NHEJ, demonstrating that ATM is involved in the
protection and repair but not the extensive resection of DSBs near telomeres (PLoS Genetics 9:e1003386,
2013). Our current studies show that the inhibition of MRE11 nuclease activity with Mirin and the
knockdown of expression of Apollo with shRNA results in an additive decrease in large deletions and GCRs
at both interstitial and telomeric DSBs, although the increase in much greater near telomeres. The
150 | P a g e
1
risk head and neck cancers after post-operative radio therapy. Narasimha Kumar Karanam, PhD ; Liang-
2
1
1
2
Hao Ding, PhD ; Uma Giri, PhD ; John V. Heymach, MD PhD ; and Michael D. Story, PhD, UT Southwestern
Medical Center, Dallas, TX and MD Anderson Cancer Center, Houston, TX
2
1
The use of miRNA as a biomarker has been proposed in a number of biological and medical
conditions such as growth and development, cancer development and metastasis, the cellular stress
response, and the radioresponse of cells and tissues. The advantages of miRNA as a biomarker lies in its
stability in tissues as well as body fluids, hence the potential for non-invasive diagnosis and prognosis. In
this study, we performed miRNA expression profiling using tumor specimens from 112 head and neck
cancer patients considered at high risk for recurrence after treatment with post-operative radiotherapy.
Specimens were triaged based upon patient outcome: no evidence of disease (NED), local recurrence (LR),
and distant metastasis (DM). Total RNA was extracted using a modified protocol of the Qiagen RNeasy
Plus kit which retains small RNA fractions. Labeled cRNAs were hybridized using the Exiqon whole genome
miRNA chip V17. Comparisons of miRNA expression between DM and NED specimens using a two-sample
t test identified 94 miRNAs were differentially expressed with statistical significance (p < 0.05), whereas
45 miRNAs were differentially expressed with statistical significance (p < 0.05) between LR and NED
specimens. Our hypothesis is; differentially expressed miRNA in the LR group would be associated with
clinical radioresistance and could serve as therapeutic targets. Amongst the miRNAs that were
significantly differentially expressed between LR and NED, we focused on miR-125a-5p as it had been
identified as a negative prognostic indicator in gastric cancers, and it targeted the ERBB2 gene. Down-
regulation of ERBB2 has been shown to result in inhibition of tumor growth and radiosensitivity in lung
and colorectal tumor cells. MiR-125a-5p expression was confirmed by qRTPCR, that is, down-regulated in
LR specimens as compared to that seen in NED specimens. Transfection of miR-125a-5p mimics into HN5
head and neck cancer cells inhibited cell proliferation and increased radiosensitivity about 30% after a 2
Gy dose, whereas miR-125-5p inhibitors increased cell proliferation and increased radio resistance about
90% after a 2 Gy dose. We are expanding our findings in a larger series of head and neck cancer cells and
confirming the regulation of ERBB2 by miR-125a-5p.
(PS2-29) The role of ATM, Apollo and MRE11 in the sensitivity of telomeric regions to DNA double-
strand breaks. Keiko Muraki; Douglas Miller; John P. Murnane, Univ. of Calif., San Francisco, San Francisco,
CA
Telomeres can interfere with DNA repair, as shown by an increase in large deletions and gross
chromosome rearrangements (GCRs), and a decrease in NHEJ at telomeric DSBs. In normal somatic cells,
the sensitivity of telomeric regions to DSBs is important in ionizing radiation-induced cell senescence and
in protection against oncogene-induced replication stress. However, in cancer cells or embryonic stem
cells, we have shown that DSBs near telomeres result in genomic instability. We previously reported that
a deficiency in ATM further increases the already high frequency of large deletions at telomeric DSBs, and
further decreases the already low frequency of distal NHEJ, demonstrating that ATM is involved in the
protection and repair but not the extensive resection of DSBs near telomeres (PLoS Genetics 9:e1003386,
2013). Our current studies show that the inhibition of MRE11 nuclease activity with Mirin and the
knockdown of expression of Apollo with shRNA results in an additive decrease in large deletions and GCRs
at both interstitial and telomeric DSBs, although the increase in much greater near telomeres. The
150 | P a g e
