Molecular progressionof thyroidlesions

Molecular progressionof thyroidlesions

Dr. Peter Lakatos1st Department of Medicine

Semmelweis University, Budapest

Dr. Peter Lakatos1st Department of Medicine

Semmelweis University, Budapest

Thyroid nodules

• Population: 4-7% with thyroid nodules

• More frequent in women

• Incidence increasing with age

• Malignancy: 5-10% of cold nodules

Basic question: Which nodule will become malignant?

Thyroid cancer

• Thyroid cancer: 1.5% of all malignancies

• Incidence increasing during the last 25 years:

4.8→ 8.0 /100,000

11.7 women, 4.2 men /100,000

History

1. Characteritics of the nodule

2. Other symptomsA.) hyperthyroidism/hypothyroidism

B) compression

3. Familiy history� MEN

4. Neck irradiation

Physical examination

• Size

• Nodule consistency, multiple, solitary

• Fixed or mobile

• Cervical lymph nodes

• Quickly enlarging nodule• Stiff, fixed nodule• Swelling cervical lypmh node• Hoarse voice, swallowing difficulty• Previous neck irradiation• Age: <20 yrs or >70 yrs• Male sex• Familiy history of medullary cancer• Familiy history of MEN-2

Signs indicating increasedrisk for malignancy inpatients with thyroid nodules

• Identifying further nodules

• Determining the size of the nodules

• Looking for malignancy signs

• Guidance of FNAB

US indication in case of thyroidnodule

• hypodensity• microcalcification• hypervascularization• solitary nodule• irregular borderline• lack of halo

The more signs are noted, the higher the risk.

Risk of thyroid cancer in a solitary nodule withmicrocalcification is 70%.

US and thyroid cancer

Thyroid Imaging Reporting and Data System (TIRADS)

Horvath E et al, JCEM 2009. 94:1748-1751.

FNAB• FNAB is the most reliable test• Results in classes:

- non-diagnostic- benign- follicular lesion- suspicious for malignancy- malignant

• Limitations of FNAB:– No differentiation between benign follicular or Hürthle-

cell adenoma and malignant forms (capsular and/orvascular invasion)

– No result in 15%• False negative: < 5%

Classificationof thyroid cancers

Follicular cells• Differentiated

– Papillary 80%– Follicular 10%– Hürthle-cell 3-5%

• Non-differentiated– Anaplastic 1-2%

Parafollicular cells– Medullary 5%

Papillary cancer

• Responsible for 90% of radiation-induced cancers

• Subclassification:: microcarcinoma, intrathyroidal, extrathyroidal– Histology variants: high-cell, clear-cell, columnar,

diffuse sclerotizing

• Multiple nodules in 30-50%

• Lymphatic spreading

• Lymph mets in 20-50%

Follicular cancer• More frequent in iodine-deficient areas• Diagnosis: vascular and/or capsular invasion• Subclassification:: minimally or extremely invasive

– Vascular invasion→ more aggressive compared to capsular invasion

• Rarely multiple nodules• Hematogenous spreading• Frequent lung and bone mets.

Hürthle -cell cancer

• Frequent mets.

• Iodine uptake in less than 10%

• Frequent recurrence

• High mortality – 30% / 10 yrs

Anaplastic cancer

• Decreasing frequency

• Rapid progression >60 yrs

• Frequent mets. at the time of dg.

• Limited surgical solutions

• Irradition or chemo not effective

• Average survival: ~ 4 - 6 months

Medullar y cancer

• Origin: parafollicular C-cells

• Calcitonin and CEA elevation in the circulation(50%)

• Sporadic: 80%

• At the time of dg.: 75%met. and 20% distantmet.

Medullar y cancer• MEN IIA �

– MC (100%), pheo (40%), hyperparathyroidim (35%)– AD trait– Missense mutation in the RET protoonkogene (extracell. cytein)– Surgery recommended before 6 yrs of age

• MEN IIB �– MC (100%), pheo (50%), mucosal ganglioneuromas (100%),

Marfan sy– AD trait– Missense mutation in the RET tirozine-kinase domain– Recommended surgery in infant age

• Familial MC

Differentiated thyroid cancers

● PTC and FTC comprise 90% of all thyroid malignancies.● PTC is the most common histological type of all thyroid malignancies (60-

80%). Somatic mutations are found in more than 40-70% of papillary carcinoma cases.

● FTC is the second most common histological type with a frequency is 10-15%. Mutation is present in 30-50%.

Benvenga, S., Horm Metab Res, 2008. 40(5): p. 323-8.Woodruff, S.L., et al., Am J Surg, 2010. 200(4): p. 462-6.Schlumberger, M., Ann Endocrinol (Paris), 2007. 68(2-3): p. 120-8.Cheng, S.P., et al., Langenbecks Arch Surg, 2008. 393(5): p. 729-32.

Albarel F, Conte-Devolx B, Oliver C.,Ann Endocrinol (Paris). 2012. Apr. 13. [Epub aheadof print]

Genetic alterations and PTC

PTC frequently carries- BRAF (v-raf murine sarcoma viral oncogene homolog B1) - RET/PTC (RET tyrosine-kinase protooncogene / papillary thyroid

carcinoma) - RAS (rat sarcoma viral oncogene homolog) mutations

BRAF mutation has been associated with more aggressive tumor behavior:- extrathyroidal extension- lymph node involvement- resistance to radioactive iodine- tumor recurrence

BRAF gene mutation is an oncogenic mutation influencing the mitogen-activated protein kinase (MAPK) signaling pathway.

THERAPEUTIC INTERVENTION!

Targeted therapy in the BRAF pathway

Cantwell-Dorris ER et al, Mol Cancer Ther; 2011. 10(3) DOI:10.1158/1535-7163.MCT-10-0799

Albarel F, Conte-Devolx B, Oliver C.,Ann Endocrinol (Paris). 2012. Apr. 13. [Epub aheadof print]

Genetic alterations and FTCFTC is prone to have

- RAS- RET/PTC - PAX8/PPAR-gamma mutations

These alterations have been identified in approximately 30-80 % of the tumors.

RAS family activates three cascades: - MAP-kinase-pathway- phosphatidilinozitol-3-kinase/protein-kinase-b (PI3K/AKT)

apoptotic-pathway- adhesive and migration affect line

The altered RAS proteins bind with major affinity to a GTP and activate GTPase effect, thus these proteins maintain constitutive activation of proliferation in follicular cells. Therefore genomic instability and increased proliferative potential will be the consequence in these cells.

Albarel F, Conte-Devolx B, Oliver C.,Ann Endocrinol (Paris). 2012. Apr. 13. [Epub aheadof print]

More genetic alterations recognizedRET proto-oncogene is a tyrosine-kinase transmembrane receptor and encoded on chromosome X. Occurence of frequent alterations in sporadic papillary cancer has been established.

- RET/PTC1 fusion protein is associated with less malignancy- RET/PTC2 is infrequent- RET/PTC3 marks more aggressive tumors and exhibits bad prognosis.

PAX8 (paired box 8) gene is encoding a transcription factor that has a role in tissue differentiation in embryonic age. The PAX8/PPAR-gamma-1 rearrangement may appear in follicular thyroid cancer.

Thyroid receptors as well as micro-RNAs might also contribute to the pathological processes resulting in malignant transformation of the thyroid.

THERAPEUTIC INTERVENTION!

Kim WG, Cheng S, Biochim. Biophys. Acta (2012), doi:10.1016/j.bbagen.2012.04.002

Thyroid carcinogenesis

de la Chapelle A, Jazdzewski K., JCEM 2011. 96:3326-36.)

Regulation of PTEN/PI3K/AKT pathway by microRNAs

Gene Cancer type Prevalence (%)

BRAF PTC 45

not well-differentiated cancer 20

anaplastic carcinoma 20

RAS anaplastic carcinoma 50

FTC 45

not well-differentiated cancer 35

PTC 10

RET medullary carcinoma 50-95

RET/PTC PTC 20

PAX8-PPARγ FTC 35

TP53 anaplastic carcinoma 70

not well-differentiated cancer 20

Genetic alterations inthyroid cancers

Age

men (n=40) 52.2 ± 13.2

women (n=85) 51.5 ± 14.2

number of DNA samples BRAF HRAS KRAS NRAS

papillary cc. 106 40 (37.7%) 1 (0.9%) 1 (0.9%) 1 (0.9%)

follicular cc. 12 2 (16.7%) 1 (8.3%) 0 3 (25.0%)

other cc.* 7 0 0 0 1 (14.3%)

normal tissue 121 0 0 0 0

summ 246

number of RNA samples

RET/PTC1 RET/PTC3

papillary cc. 59 5 (8.5%) 1 (1.7%)

follicular cc. 12 0 0

other cc.* 7 0 0

normal tissue 78 0 0

summ 156

*: anaplastic thyroid carcinomas, follicular adenomas and follicular neoplasia

Genetic alterations in thyroid cancers in Hungary

Balla B et al, Thyroid 2011. 21(4):459-460.

CYP24A1 has beenidentified in PTC

• CYP24A1 is the calcitriol-neutralizing enzyme• Increased expression has been shown in colon cc and liver cc

Changes of relative CYP24A1 gene expression in 51 human papillary tumors vs.

control thyroid tissues.

Patients’ number

CYP24A1 mRNA expression was markedly increased in 31 cases (61%) of the examined papillary cancer compared with that

of normal thyroid tissue, sometimes reaching over 106-fold elevation. We measured lower CYP24A1 transcription level in

12 tumor tissues (23.5%). There was no variation in CYP24A1 gene expression between 2 tumor and healthy control tissue,

and CYP24A1 specific mRNA was not detected in the thyroid samples of 6 patients.

Component143210-1-2-3-4

Com

pone

nt2

4

3

2

1

0

-1

-2

-3

-4

20

30

38

39

26

1

31

11

6

8

40

19 18

21

17

10

13

12

32

34

149

37

41

25

22

29

5

42

35

4

23

333

272

7

28

24

36

16 44

15 43

Tumorlocalization 1

Winter

Lymph node metastasis

PTC oncocyter variant

CCDC6/RET

Hashimoto

Hyperthyreosis

CYP24A1 RATIOKRAS

Vascularization

Spring

Hypothyreosis

No other thyroid disease

DIRECTION

BRAFTumor size Hyperplasia

Age

ELE1/RET Tumor localization 2

Lymphocytas thyreoiditisSex

No PTC subtype/variation

Fall

HRAS

Tumor localization 3

Struma

Summer

PTC follicular variant

PTC hyalinized variant

Principal component analysis

Component143210-1-2-3-4

Com

pone

nt2

4

3

2

1

0

-1

-2

-3

-4

Tumorlocalization 1

Winter

Lymph node metastasis

PTC oncocyter variant

CCDC6/RET

Hashimoto

Hyperthyreosis

CYP24A1 RATIOKRAS

Vascularization

Spring

Hypothyreosis

No other thyroid disease

DIRECTION

BRAFTumor size Hyperplasia

Age

ELE1/RETTumor localization 2

Lymphocytas thyreoiditisSex

No PTC subtype/variant

Fall

HRAS

Tumor localization 3

Struma

Summer

PTC follicular variant

PTC hyalinized variant

Principal component analysis

HMGA2 (high mobility group AT-hook 2): RAS-regulatedcell proliferationpancreatic ductal adenocarcinoma, papillary serous carcinoma

MRC2 (mannose receptor, C type 2, also known as Endo 180)tumor cell migration

SFN/14-3-3r (stratifin)tumor suppressor genelung cancer, squamous cell carcinoma, pancreatic cancer,endometrial carcinoma, cervical cancer, colorectal carcinoma

Prasad NB et al, Thyroid, 2012. 22(3):275-284.

Three-Gene Molecular Diagnostic Modelfor Thyroid Cancer

Specificity: 100%Sensitivity: 80%

Follow-up study

● FNAB samples collected● Planned: 800, accomplished to date: 710● At least 3 yrs of follow-up planned● Frequency of genetic alterations, incidence of malignancies recorded● Can genetic alterations predict malignancy?

Genetic alteration Frequency

BRAF 4

NRAS 1

HRAS 7

KRAS 0

RET/PTC3 1

One year follow-up of 231 subjects

13

Malignant samples (cytology)

Follicular adenoma (4)

PTC 14

Medullary 1

15 (19)

malignant non-malignant

+ -

Geneticalteration

+ 5 8

- 10 223

231

4 BRAF and 1 RET/PTC3

Melck AL, Yip L, HEAD & NECK—2011, DOI 10.1002/hed.21818.

University of Pittsburgh’s current algorithm for the evaluation of thyroid nodules

Contribution of molecular information to thyroiddiagnostics and therapy

● Additional information to FNAB or histology

● Individual treatment options (e.g. thyrosine kinase inhibition)

● Prediction of malignancy in thyroid nodules

Thank you foryour attention!