Research Presentation Jason M. Leibowitz, MD June 25, 2009 Preceptor: Marcia S. Brose, MD PhD Jason...

Research PresentationResearch Presentation

Jason M. Leibowitz, MD

June 25, 2009

Preceptor: Marcia S. Brose, MD PhD

Jason M. Leibowitz, MD

June 25, 2009

Preceptor: Marcia S. Brose, MD PhD

Otorhinolaryngology: Head and Neck Surgery at PENN Excellence in Patient Care, Education and Research since 1870

OverviewOverview

Background Hypothesis Methods Results Discussion Conclusions & Future Directions

Thyroid Cancer in the United StatesThyroid Cancer in the United States

Thyroid cancer is the most

common endocrine neoplasm.

Thyroid cancer will be diagnosed

in 33,550 individuals (8070 men

and 25,480 women) this year.

From 1997-2004 incidence of

thyroid cancer increased by 6.2%

mostly due to increased detection.

From 1985 to 2004 mortality rate

increased by 0.3% a year.

Thyroid cancer is the most

common endocrine neoplasm.

Thyroid cancer will be diagnosed

in 33,550 individuals (8070 men

and 25,480 women) this year.

From 1997-2004 incidence of

thyroid cancer increased by 6.2%

mostly due to increased detection.

From 1985 to 2004 mortality rate

increased by 0.3% a year.

RAI-Refractory DiseaseRAI-Refractory Disease

25-50% of metastatic thyroid cancers lose ability to take up Iodine.

Iodine Uptake inversely correlates with survival. This is attributed to down regulation of the Na+/I-

Symporter (NIS). Limited treatment options for unresectable

thyroid cancer refractory to RAI.

25-50% of metastatic thyroid cancers lose ability to take up Iodine.

Iodine Uptake inversely correlates with survival. This is attributed to down regulation of the Na+/I-

Symporter (NIS). Limited treatment options for unresectable

thyroid cancer refractory to RAI.

Molecular Changes in Thyroid Cancer

Molecular Pathway involved in Thyroid Cancer

Activation of MAPK pathway Oncogenic activation

of this pathway in 70% of all thyroid cancers.

BRAF is a serine threonine kinase

Activation of MAPK pathway Oncogenic activation

of this pathway in 70% of all thyroid cancers.

BRAF is a serine threonine kinase

Xing, 2007.

BRAF V600E in Thyroid CancerBRAF V600E in Thyroid Cancer

2003: The BRAF V600E mutation is the most

common genetic alteration in thyroid cancer,

occurring in about 45% of sporadic papillary

thyroid cancers (PTCs).

2003: The BRAF V600E mutation is the most

common genetic alteration in thyroid cancer,

occurring in about 45% of sporadic papillary

thyroid cancers (PTCs).

BRAF V600EBRAF V600E Point mutation in 40-45% of

PTC Upregulation of MMP, VEGF -->

invasion, angiogenesis Silencing of tumor suppressive

genes, genes involved in iodine transport

BRAF mutation associated with multiple negative prognostic indicators.

Point mutation in 40-45% of PTC Upregulation of MMP, VEGF -->

invasion, angiogenesis Silencing of tumor suppressive

genes, genes involved in iodine transport

BRAF mutation associated with multiple negative prognostic indicators.

RASRAS

Family of small G-proteins involved in transduction of cellular signals from the cell membrane.

Mutations in RAS gene lead to inappropriate activation with constitutively activated downstream pathways and also promote chromosomal instability.

20% FTC contain a RAS mutation RAS mutations may correlate with aggressive behavior (tumor dedifferentiation and poorer prognosis).

Family of small G-proteins involved in transduction of cellular signals from the cell membrane.

Mutations in RAS gene lead to inappropriate activation with constitutively activated downstream pathways and also promote chromosomal instability.

20% FTC contain a RAS mutation RAS mutations may correlate with aggressive behavior (tumor dedifferentiation and poorer prognosis).

Targeted Therapy in Thyroid cancer

Loss of differentiation (inability to trap RAI), unresectable lesion, leads to poor prognosis

BRAF inhibitors BAY 43-9006 (Sorafenib) Multikinase inhibitor

Loss of differentiation (inability to trap RAI), unresectable lesion, leads to poor prognosis

BRAF inhibitors BAY 43-9006 (Sorafenib) Multikinase inhibitor

SorafenibSorafenib

Orally active multikinase inhibitor (study dose 400mg BID).

Monoclonal antibody with multiple targets including BRAF, VEGFR1, VEGFR2.

Blocks tumor cell proliferation and angiogenesis.

FDA approved for treatment of RCC and hepatocellular carcinoma.

Orally active multikinase inhibitor (study dose 400mg BID).

Monoclonal antibody with multiple targets including BRAF, VEGFR1, VEGFR2.

Blocks tumor cell proliferation and angiogenesis.

FDA approved for treatment of RCC and hepatocellular carcinoma.

Targeted Therapy and GenotypeTargeted Therapy and Genotype

K-RAS gene mutation and metastatic colorectal carcinoma. Recent results from Phase II & III clinical trials demonstrate that

patients with metastatic colorectal cancer benefit from anti-EGFR therapy.

Patients with K-RAS mutation in codon 12 & 13 should not receive anti-EGFR therapy since they do not receive any benefit.

EGFR and non-small cell lung cancer: Epithelial growth factor receptor 10% mutated in NSCLC EGFR mutations are predictors of TKIs responsiveness and may

show a long lasting response to TKIs EXON 19 Deletion respond better to TKIs.

K-RAS gene mutation and metastatic colorectal carcinoma. Recent results from Phase II & III clinical trials demonstrate that

patients with metastatic colorectal cancer benefit from anti-EGFR therapy.

Patients with K-RAS mutation in codon 12 & 13 should not receive anti-EGFR therapy since they do not receive any benefit.

EGFR and non-small cell lung cancer: Epithelial growth factor receptor 10% mutated in NSCLC EGFR mutations are predictors of TKIs responsiveness and may

show a long lasting response to TKIs EXON 19 Deletion respond better to TKIs.

Prior DataPrior Data

84 weeks

Papillary vs. FollicularPapillary vs. Follicular

P<0.095FTC = 19

PTC= 24

Prior DataPrior Data

Conclusions from prior data: Improved PFS with Sorafenib. Improved PFS of FTC treated with Sorafenib

when compared to PTC.

Conclusions from prior data: Improved PFS with Sorafenib. Improved PFS of FTC treated with Sorafenib

when compared to PTC.

OverviewOverview

HypothesisHypothesis

There are specific genotypes (i.e. BRAF V600E, RAS mutations) that predict favorable response to targeted therapy (Sorafenib).

Null HypothesisNull Hypothesis

Specific genetic mutations do not predict response to targeted therapy in thyroid cancer.

OverviewOverview

Research PlanResearch Plan Tissue samples collected from patients with

treatment-resistant thyroid cancer with long term follow-up (approximately 30 patients).

All patients received targeted therapy (Sorafenib). Samples with WDTC analyzed for mutations in

BRAF and RAS genes when available: BRAF - V600E RAS - Exon 12, 13, 61

Tissue samples collected from patients with treatment-resistant thyroid cancer with long term follow-up (approximately 30 patients).

All patients received targeted therapy (Sorafenib). Samples with WDTC analyzed for mutations in

BRAF and RAS genes when available: BRAF - V600E RAS - Exon 12, 13, 61

RESULTS

Sequence OutputSequence Output Computer program interprets

data and produces an electropherogram, (aka trace)

Each peak represents a base: A = Adenosine T = Thymine C = Cytosine G = Guanine N = Reading cannot be

determined

Computer program interprets data and produces an electropherogram, (aka trace)

Each peak represents a base: A = Adenosine T = Thymine C = Cytosine G = Guanine N = Reading cannot be

determined

OverviewOverview

Results of Stage 1 AnalysisResults of Stage 1 Analysis•N= 30 •M = F = 15•PTC=17, FTC= 9, Other (ATC/PD, MTC): 4•Samples analyzed for BRAF mutation:

• 23/30 (76.6%): samples analyzed for BRAF mutation• 4/30 (13%): definite genotype but questioned due to phenotype

(ATC/PD, MTC)• 2/30 (6%): unable to amplify DNA despite multiple PCR attempts• 1/30 (3%): pending analysis

•18/30 samples analyzed for RAS mutation, all WT copies of the gene

•N= 30 •M = F = 15•PTC=17, FTC= 9, Other (ATC/PD, MTC): 4•Samples analyzed for BRAF mutation:

• 23/30 (76.6%): samples analyzed for BRAF mutation• 4/30 (13%): definite genotype but questioned due to phenotype

(ATC/PD, MTC)• 2/30 (6%): unable to amplify DNA despite multiple PCR attempts• 1/30 (3%): pending analysis

•18/30 samples analyzed for RAS mutation, all WT copies of the gene

Results of Stage 1 AnalysisResults of Stage 1 Analysis

N=22 (interim analysis) 13 WT BRAF 9 BRAF V600E 16 PTC

9 WT BRAF, 7 V600E

6 FTC 4 WT BRAF, 2 V600E

N=22 (interim analysis) 13 WT BRAF 9 BRAF V600E 16 PTC

9 WT BRAF, 7 V600E

6 FTC 4 WT BRAF, 2 V600E

BRAF V600EBRAF V600E

P<0.02

N=13 (WT=8,

V600E=5)

Updated geneticsUpdated genetics In our expanded

analysis to 22 pts with WDTC, the effect is no longer significant but the trend exists.

We are further investigating BRAF copy number in these patients

In our expanded analysis to 22 pts with WDTC, the effect is no longer significant but the trend exists.

N =22WT = 13BRAF V600E = 9

OverviewOverview

BRAFV600E Correlates with worse SurvivalBRAFV600E Correlates with worse Survival

Elisei et. al, J Clin Endocrinol Metab, October 2008, 93(10):3943–3949

BRAFV600E Correlates with worse SurvivalState of the mutation in PTC, 10/2008

THE BRAF connectionTHE BRAF connection

Ciampi et al. 2005

Updated geneticsUpdated genetics In our expanded

analysis to 22 pts with WDTC, the effect is no longer significant but the trend exists.

In our expanded analysis to 22 pts with WDTC, the effect is no longer significant but the trend exists.

N =22WT = 13BRAF V600E = 9

BRAF (red) x 3

7 centromere (green) x 3

BRAF x4

7 centromere x4

4 copies each

3 copies each

THE BRAF connection! Positive Predictor!

Ciampi et al, 2005.

Future DirectionsFuture Directions

Completion of genotyping analysis of all patients

Evaluation of copy number gains in WDTC Hypothesis: Copy number gain accounts for

improved survival in FTC treated with Sorafenib

Null: Copy number gain does not influence survival in FTC

Completion of genotyping analysis of all patients

Evaluation of copy number gains in WDTC Hypothesis: Copy number gain accounts for

improved survival in FTC treated with Sorafenib

Null: Copy number gain does not influence survival in FTC

Selected SourcesSelected Sources

Ciampi R, Zhu Z, Nikiforov YE. BRAF copy number gain in thyroid tumors detected by fluorescence in situ hybridization. Endocrine Pathology 2005; 16(2): 99-105.

Ciampi R, Nikiforov YE. Alterations of the BRAF gene in thyroid tumors. Endocrine Pathology 2005; 16:3): 163-171.

Gupta-Abramson V, Troxel AB, Nellore A, et al. Phase II Trial of Sorafenib in Advanced Thyroid Cancer. Journal Clin Onc 2008; 26 (29): 4714-4719.

Kundra P, Burman KD. Thyroid Cancer Molecular Signaling Pathways and Use of Targeted Therapy. Endoc Metab Clin N Am 2007;36: 839-853

Murer B. Targeted Therapy in Non-Small Cell Lung Cancer. Arch Path Lab Med. 2008; 132: 1573-1575.

Nikiforov YE. Thyroid Carcinoma: Molecular Pathways and Therapeutic targets. Modern Pathology 2008; 21: S37-S43.

Vasko V, Ferrand M, Cristofaro JD et al. Specific Pattern of RAS Oncogene Mutations in Follicular Thyroid Tumors. J. Clin Endocrin. & Metab. 2003; 88(6):2745-2752.

Xing M. BRAF Mutation in Papillary Thyroid Cancer: Pathogenic Role, Molecular Basis, and Clinical Implication. End Rev 2007; 28(7): 742-762.