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Oral Papers
the 3’UTR of messenger RNA (mRNA), causing translational repression
or degradation. To better understand the potential role of miRNA in
HNSCC carcinogenesis, miRNA-21 specific gene targets were identified
and their expression signatures were assessed utilizing the power of
microarray technology and bioinformatics miRNA target prediction
tools. Design: Whole genome mRNA expression was analyzed by
microarrays in HNSCC tumors, normal mucosa and a normal oral
keratinocyte cell line (NOK-SI) transiently overexpressing miRNA-21. A
candidate list of miRNA-21 gene targets was generated by identifying
common downregulated mRNA expression signatures in HNSCC
tumors and the miRNA-21 overexpressing normal oral keratinocyte cell
line. The candidate target gene list was further narrowed by evaluating
each 3’UTR region with bioinformatics tools for miRNA-21 specificity.
Expression levels of miRNA-21 gene targets were determined by
reverse transcription quantitative polymerase chain reaction (RTqPCR).
Specificity of miRNA-21 for each candidate gene target was
assayed utilizing the luciferase miRNA target expression vector system.
Subjects: Matched 13 HNSCC tumors and 5 normal mucosa total RNA
samples, in addition to NOK-SI cell line RNA samples after triplicate
miRNA-21 and mock lentiviral vector transient transfections were used
for mRNA microarray analysis. Results: Microarray analysis combined
with bioinformatics tools for miRNA-21 gene target prediction generated
a candidate gene list, including Clusterin, Basonuclin 2, and Desmin
(FAM48a). Expression levels of miRNA-21 gene targets were significantly
lower in HNSCC tumors and miRNA-21 transfected NOK-SI cell line
when compared to normal mucosa and mock transfections by RTqPCR.
Transfection of miRNA-21 significantly suppressed a luciferasereporter
containing the 3’UTR of the miRNA-21 gene target verifying the
specificity of miRNA-21 for the candidate. Conclusions: Novel miRNA
targets can be identified and validated utilizing an integrative approach
with whole genome expression profiling and bioinformatics target
prediction methods. MiRNA-21 has several mRNA targets in HNSCC and
further study is needed to evaluate their functional effects and possible
use in detection, prognosis, and therapeutic outcome in HNSCC.
S015
AKT PHOSPHORYLATION AS A MARKER OF RESISTANCE TO
ZD6474 (VANDETANIB) IN HEAD AND NECK SQUAMOUS CELL
CARCINOMA. Genevieve A Andrews, MD, Mitchell J Frederick, PhD,
Mei Zhao, MD, David R Fooshee, Zvonimir L Milas, MD, Maria K Gule,
MD, Chad E Galer, MD, Jeffrey N Myers, MD PhD; MD Anderson Cancer
Center.
The epidermal growth factor receptor (EGFR) is an attractive target
for anti-cancer therapy, in head and neck squamous cell carcinoma
(HNSCC). Yet often HNSCC demonstrates resistance to EGFR inhibition.
Therefore, drugs that target angiogenesis by inhibiting the VEGFR
(vascular endothelial growth factor receptor) in addition to EGFR, such
as ZD6474 (Vandetanib), may be useful for treating HNSCC patients.
To better understand the mechanisms of resistance to EGFR inhibition,
as well as examine biomarkers defining EGFR inhibitor resistance, we
investigated the effect of ZD6474 in a panel of 50 HNSCC lines in vitro,
where the principal in vitro drug effects are thought to be through the
EGFR signaling pathway. The GI50 (50% growth inhibition) values for
cell lines ranged from 0.4 µM to 14 µM between the most sensitive and
most resistant lines. When sensitive HNSCC cell lines FADU and PCI-13
were treated for 30 hours with 4 µM ZD6474, there was more than a 70%
reduction in the phosphorylation of AKT on Ser427. However, there was
less than a 10% decrease in pAKT Ser427 observed for two resistant
cell lines examined, SCC61 and JHU 028. There was a parallel reduction
in constitutive pERK in the two sensitive cell lines following treatment
with ZD6474 for 30 hours. The reduction in pERK only occurred in one
of the resistant lines, SCC61. EGF-stimulated phosphorylation of the
EGFR was similarly inhibited in both resistant and sensitive cells by 4 µM
ZD6474, indicating that resistance was not due to a difference in drug
target sensitivity in these cell lines. In conclusion, our preliminary data
suggest a model wherein HNSCC cells sensitive to EGFR inhibition rely
heavily upon the EGFR pathway to activate and maintain proliferation
and survival mechanisms such as phosphorylation of AKT, whereas
resistant cells may use other receptors or signaling molecules to activate
critical downstream pathways to survive in the face of EGFR inhibition.
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S016
A BLOOD TRANSFUSION PREDICTION MODEL IN PATIENTS
UNDERGOING MAJOR HEAD & NECK SURGERY INVOLVING FREE
FLAP RECONSTRUCTION. Manish D Shah, MD MPhil FRCSC, David
P Goldstein, MD FRCSC, Stuart McClusky, MD FRCPC, Patrick Gullane,
MB FRCSC FACS FRACS Hon, Dale H Brown, MD FRCSC, Jonathan C
Irish, MD MSc FRCSC FACS, Ralph W Gilbert, MD FRCSC; University
Health Network, University of Toronto, Toronto, Canada.
Objectives: Perioperative blood transfusion is commonly required
for major head & neck procedures and carries significant risks.
Alternatives to allogenic blood transfusion are becoming available.
Pre-operative risk stratification of patients allows for appropriate
patient counseling and resource allocation. Thus, the objective of our
study was to develop a model to reliably predict the requirement for
perioperative blood transfusion in patients undergoing major head and
neck surgery involving free flap reconstruction. Methods: Data was
prospectively collected on all patients undergoing major head and
neck surgery requiring free flap reconstruction at the University Health
Network (Toronto, Canada) between 1999 and 2009. Over 800 patients
were included in the analysis. Pre-operative variables were tested for
association with the outcome of interest, perioperative transfusion.
Stepwise multivariable logistic regression modeling was carried out
to determine which pre-operative variables were best able to predict
perioperative transfusion requirement. Results: A predictive model
of perioperative blood transfusion requirement was obtained from the
logistic regression analysis. Six pre-operative variables were found to
be significant—female gender (odds ratio [OR] = 4.1), T-stage (T1/2 vs
T3/4, OR = 1.4), treatment with pre-operative chemotherapy (OR = 1.9),
cardiac comorbidity (OR = 1.8), pre-operative hemoglobin level (normal
vs low, OR = 5.6), and type of free flap reconstruction (non-osseus vs
osseus, OR = 2.3). Amongst these six variables, gender, pre-operative
hemoglobin level, and type of free flap were the strongest predictors in
the model. Using this model, we can predict the probability of a patient
requiring a perioperative blood transfusion. Conclusions: We have
developed a reliable model for predicting perioperative blood transfusion
requirements in patients undergoing major head and neck surgery
requiring free flap reconstruction. This model can be used for accurate
pre-operative risk stratification. This is essential for effective patient
counseling, use of cross and type, and use of alternatives to allogenic
blood transfusion.
S017
A CLASSIFICATION SYSTEM FOR RECONSTRUCTION OF VERTICAL
HEMIPHARYNGOLARYNGECTOMY FOR HYPOPHARYNGEAL
SQUAMOUS CELL CARCINOMA. Min-Sik Kim, MD PhD, Young-Hoon
Joo, MD, Dong-Il Sun, MD PhD, Kwang-Jae Cho, MD PhD, Jun-Ook
Park, MD; The Catholic University of Korea.
Objectives: To evaluate microvascular reconstruction of vertical
hemipharyngolaryngectomy (VHPL) defect for hypopharyngeal
squamous cell carcinoma. We classified VHPL according to the extent
of tumor resection to establish guidelines for its clinical application.
Methods: We have reviewed a 12-year experience with 32 VHPL
between 1998 and 2009. Based on our retrospective reassessment, the
classification comprised three types of VHPL according to the extent
of resection: limited VHPL (type I ), resection at the lateral border of
the conus elasticus to preserve both vocal cords (n=10); VHPL (type
II), removal of a vertical section of the thyroid cartilage through the
anterior commissure to the upper border of the cricoid cartilage with
preservation of one vocal cord (n=12); and extended VHPL (type III),
inclusion of a supraglottic laryngectomy (type IIIa) (n=6) and/or partial
cricoid cartilage resection (type IIIb) (n=4). A radial forearm free flap
that included the palmaris longus tendon was used for reconstruction
in 31 patients, and an anterolateral thigh flap was used in one patient.
Results: There was no perioperative mortality, and there was a 100%
free flap survival rate. Two (6.3%) patients developed a postoperative
pharyngocutaneous fistula and 2 (6.3%) patients had a glottic stenosis
develop during the postoperative period. Oral re-alimentation was
achieved within a mean of 33 days postoperatively. Twenty eight (88%)
patients could achieve all of their nutritional needs orally; however, four
(12%) patients needed the assistance of a PEG tube with type II and IIIa
defect. Tracheotomy weaning was achieved within a mean of 31.9 days
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