Roma 26-27 Giugno 2015

Multidisciplinarietà e Biologia Molecolare applicate alla pratica

Cli i O l i ' t ità d t i t ttiClinica Oncologica: un'opportunità ed un vantaggio per tutti

Quale messaggio la Biologia Molecolare gg gtrasferisce alla pratica clinica

Alvaro Leone

Anatomia patologica p g

Ospedale San Camillo-Forlanini Roma

Direttore: Dr.ssa Lucia Grillo

After an initial response nearly all EGFR-mutated patients eventually develop resistance to reversible EGFR-TKIs

D l 19Del 19

L858R

Shigematsu H et al. JNCI J Natl Cancer Inst 2005;97:339-346

Journal of the National Cancer Institute, Vol. 97, No. 5, © Oxford University Press 2005, all rights reserved.

NEJM 352;8 february 24, 2005

…The DNA sequence of the EGFR gene in tumor biopsy i l l d h f d ispecimen at relapse revealed the presence of a second point

mutation, resulting in threonine-to-methionine amino acid h i i 790 f EGFRchange at position 790 of EGFR…

TKIs : How they work

ATP

TKIs : How they work

ATPGEFITINIB/ERLOTINIB

TKIs : How they work

E 20 T790MExon 20 T790M

TKIs : How they work

E 20 T790MExon 20 T790MTKI

The Biology of resistant disease

Synergistic transforming activity of T790M in ciswith EGFR mutations

T790M/DelRas

Del

T790M

Godin-Heymann et al. Cancer Res. 2007 august 1; 67(15):7319-7326

Postprogression survival (A) and time from progression until new metastasis (B) depending on

Clinical behaviour of resistant disease

Postprogression survival (A) and time from progression until new metastasis (B), depending on T790M mutation status.

Oxnard G R et al. Clin Cancer Res 2011;17:1616-1622

©2011 by American Association for Cancer Research

Highly Sensitive Detection of EGFR T790M Mutation Using Colony Hybridization PredictsFavorable Prognosis of Patients with Lung Cancer Harboring Activating EGFR Mutation

Fujita y et al. J Thoracic oncology 2012; 7:1640-44

EGFR Mutation and Resistance of Non–Small-Cell Lung Cancer to Gefitinib

Susumu Kobayashi et al. N England J Med 2005; 352:786-792To obtain further evidence of the presence of a second mutation, cDNA was generated from RNA isolated from the paraffin block of the biopsy specimen obtained at the time offrom RNA isolated from the paraffin block of the biopsy specimen obtained at the time ofrelapse. The cDNA was amplified, and the C-to-T base-pair change was confirmed in 14 of 40 subclones. Interestingly, the C-to-T base-pair change was consistentlyobserved with either wild type or delL747 S752 sequences suggesting that the mutationobserved with either wild-type or delL747–S752 sequences, suggesting that the mutationis either biallelic or that the tumor has two distinct populations of cells.

Peptide nucleic acid–locked nucleic acid polymerase chain reaction clamp-based detection test for gefitinib-refractory T790M epidermal growth factor receptor mutation

1. Hitoshi Miyazawa et al. Cancer Sci 2008; 99:595-600

Next we subcloned the RT-PCR product into a plasmid, picked up 13 clones andNext we subcloned the RT PCR product into a plasmid, picked up 13 clones and sequenced them. Two out of the 13 clones had both the T790M and L858R mutations;the remaining 11 had only the L858R mutation

The Heterogeneity IssueThe Heterogeneity Issue

Does T790M Disappear? Successful Gefitinib Rechallenge After T790M Disappearance in a Patient With EGFR-Mutant Non–Small-Cell Lung Cancer

Hata, Akito MD; Katakami, Nobuyuki MD, PhD; Kaji, Reiko MD; Fujita, Shiro MD, PhD; Imai, Yukihiro MD, PhD

J Thoracic Oncology Volume 8 (3) march 2013

Lung LiverLesson from patients: Is T790M always present in all sites of diseasprogression?

B

g

A

Pre-TKI

C DC D

After 2 cycles

E F

After 4 cycles

Pre-TKI Lung After 4 cycles (Lung) After 4 cycles (Liver)

FISH for EGFR A D G

T>G L858R T>G L858R

Ex 21 L858R

T>G L858R T>G L858R T>G L858R

B E H

Ex 20 T790MC>T T790Mwild-type wild-typeC F I

Fig. 3

Consideration:

Rebiopsy of the lung lesion would have beenmisleadingmisleading

The accurate molecular comprehension of systemicThe accurate molecular comprehension of systemic disease is paramount to design the best therapy approachapproach

59% i T790M59% in T790M pos

29% in T790M neg

61% in T790M pos

21% i T790M21% in T790M neg

Medicine Volume 94, Number 21, May 2015

This systematic review included 25 studies with 2605 patients.

The pooled overall sensitivity specificity and concordance rate were 0 61 0 90 and 0 79The pooled overall sensitivity, specificity, and concordance rate were 0.61, 0.90, and 0.79

Serum showed lower sensitivity (0.56 vs 0.65) but higher specificity (0.95 vs 0.85) and higher concordance (0 86 vs 0 74) than plasmaconcordance (0.86 vs 0.74) than plasma.

EGFR mutations (exon 19 or 21) in blood were significantly associated with objective response(RR: 4 08; 95% confidence interval [CI] 2 48–6 70) PFS (HR: 0 72; 95% CI 0 64–0 80) and OS(RR: 4.08; 95% confidence interval [CI] 2.48 6.70), PFS (HR: 0.72; 95% CI 0.64 0.80), and OS(HR: 0.71; 95% CI 0.50–0.99).

Blood in particular serum is a good substitute when tumor tissue is absent or insufficient forBlood, in particular serum, is a good substitute when tumor tissue is absent or insufficient for testing EGFR mutations to guide EGFR TKIs treatment in patients with NSCLC. EGFR mutation positivity in blood could be used to recommend EGFR TKIs treatment, but the absence of blood positivity should not necessarily be construed with confirmed negativity.

Cellule Resistenti ai TKI di terza generazione sono sensibili ai TKI di prima generazione

del/T790M/C797Sdel/C797Sdel

Rappresentazione schematica delle Mutazioni di resistenza ed eventuali

Possibilità di terapiaPossibilità di terapia

The real success of Molecular Biology has been the change of cancer patient's treatmentchange of cancer patient's treatment

Pre- Molecular-era Molecular-era

• PatientSurgeon

• Patient• Surgeon• Surgeon

PathologistHi t l

Su geo• Pathologist• Source of information• Histology

• Diagnosis

• Source of information • Oncologist

P th l i t• Oncologist • Pathologist

Molecular biologistMolecular biologist