La ricerca traslazionale e
le ricadute nella pratica clinica
Nicola Normanno
ISTITUTO NAZIONALE PER LO STUDIO E LA CURA DEI TUMORI FONDAZIONE G. Pascale – NAPOLI
SC Biologia Cellulare e Bioterapie
CENTRO RICERCHE ONCOLOGICHE MERCOGLIANO (AV)
Laboratorio di Farmacogenomica
Molecular alterations in cancer cells offer potential for therapeutic intervention with
"target-based agents"
Hanahan & Weinberg Cell 2011
The efficacy of targeted therapy depends on
TUMOR HETEROGENEITY
Burrell Nature 2013
Meric-Bernstam JCO 2013
Target-based agents + predictive biomarkers:
PERSONALIZED MEDICINE
Predictive biomarkers and personalized medicine
• Biomarkers that are associated with
response to drugs (positive selection)
– EGFR mutations and ALK rearrangements in NSCLC
– BRAF mutations in melanoma
– ERBB2 gene
amplification in
breast/gastric cancer
• Biomarkers that are associated with
resistance to drugs (negative selection)
– RAS mutations and resistance to EGFR
monoclonal antibodies
in CRC
A patient story in 2010
• On November 2010, a 41-year-old man, former light smoker (5 pack- years), was admitted to our Institution
• Critical clinical conditions: massive pericardial effusion, disseminated intravascular coagulation, renal and hepatic failure (ECOG PS 4).
• CT scan showed two lung lesions in the upper right lobe (24 and 21 mm), right pleural effusion, a suspected metastatic lesion of the right ventricle of the heart (16 x 22 mm), confirmed by cardiac ultrasonography,
thromboembolic defects of the right pulmonary artery, multiple hepatic lesions (20 mm), thrombosis of the portal system with intrahepatic
cavernous angiomas, ascites, multiple bone metastases (sternum, cervical, dorsal and lumbar spine, pelvis).
• The cytologic evaluation of the pericardial effusion was positive for lung adenocarcinoma.
• Mutational analysis of the EGFR was requested on a cytologic specimen containing only 20 cells.
Morabito JTO 2012
EGFR mutations and NSCLC
Lynch NEJM 2004; Paez Science 2004; Pao PNAS 2004; Sequist JCO 2007; Mok NEJM 2009
EGFR mutational analysis of CTCs,
serum and plasma by using PNA-clamp
168bp DNA DEL
35 CTCs
STANDARD
0 CTCs
2 CTCs
183bp DNA WT
168bp DNA DEL
STANDARD
183bp DNA WT
168bp DNA STANDARD DEL
PLASMA - PNA
+ PNA
STANDARD STANDARD
183bp DNA WT
168bp DNA DEL
SERUM - PNA
+ PNA
*: obtained from the same patient at different time points during the treatment
Samples analysed
Tumor EGFR status
Source of tumor DNA
EGFR mutations Ex 19 Del L858R
1*
c.2235_2249del15 p.E746_A750del
CTC (35) Mut WT
Serum Mut WT
Plasma Mut WT
2* c.2235_2249del15
p.E746_A750del CTC (0) WT WT
3*
c.2235_2249del15 p.E746_A750del
CTC (2) Mut WT
Serum Mut WT
Plasma Mut WT
Morabito JTO 2012
December 2010 March 2011 July 2011
• On December 15, we started therapy with gefitinib 250 mg/day, with a progressive and dramatic improvement of clinical conditions. After two weeks of treatment, renal and hepatic functions were normalized
• The PET scan after 3 months of gefitinib showed only a minimal tracer accumulation at the basis of the right lung (SUV 3.3), but became
negative after 7 months of therapy, indicating a complete response
• Response to gefitinib was mantained for 11 months, until November 2011 when an epatic progression occurred
• The patient died on March 2012
Morabito JTO 2012
Genomic alterations affecting actionable signaling pathways
Garraway JCO 2013
Genomics-Driven Oncology
Garraway JCO 2013
Clinical trial designs utilizing molecular
profiling
Kummar JNCI 2015
Challenges in genomics-driven oncology
• Alterations of “actionable” oncogenic pathways are not always predictive of response to targeted agents
• Different molecular alterations of the same oncogene may not be equivalent
• Intra-tumor heterogeneity may affect response to targeted agents
• The molecular profile of solid tumors may significantly
change following treatment with target based agents
BRAF mutations in melanoma, CRC and NSCLC
Cancer Frequency Prognostic Predictive§ %V600E
Melanoma 50% Y/N Y 90%
CRC 10% Y N 90%
NSCLC 3% Y* Y 50%
*only V600E
§for response to cIass I RAF kinase inhibitors
Challenges in genomics-driven oncology
• Alterations of “actionable” oncogenic pathways are not always predictive of response to targeted agents
• Different molecular alterations of the same oncogene may not be equivalent
• Intra-tumor heterogeneity may affect response to targeted agents
• The molecular profile of solid tumors may significantly
change following treatment with target based agents
• EGFR mAbs have been approved by the EMA for the treatment of patients with
wild-type RAS mCRC1,2
• Mutational status should be determined by an experienced laboratory using validated test methods for detection of
KRAS (exons 2, 3 and 4) and NRAS (exons 2, 3 and 4) mutations prior to treatment with cetuximab or panitumumab1,2
1. Vectibix® (panitumumab) Summary of Product Characteristics, Amgen;
2. Erbitux® (cetuximab) Summary of Product Characteristics, Merck Serono;
3. Normanno N, et al. Nat Rev Clin Oncol 2009;6:519−27.
EGFR mAbs in CRC
RAS mediated intracellular signal transduction pathways3
G13D KRAS mutations and outcome
De Roock JAMA 2010
Challenges in genomics-driven oncology
• Alterations of “actionable” oncogenic pathways are not always predictive of response to targeted agents
• Different molecular alterations of the same oncogene may not be equivalent
• Intra-tumor heterogeneity may affect response to targeted agents
• The molecular profile of solid tumors may significantly
change following treatment with target based agents
The efficacy of targeted therapy depends on
TUMOR HETEROGENEITY
Burrell Nature 2013
0,00 20,00 40,00 60,00 80,00 100,00 120,00 140,00
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 HS SCORE
PIK3CA
0,00 20,00 40,00 60,00 80,00 100,00 120,00 140,00 160,00
1 2 3 4 5 6 7 8 9 10 11 12 13
HS SCORE
NRAS
0,00 20,00 40,00 60,00 80,00 100,00 120,00 140,00
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
HS SCORE
BRAF
* *
* *
* *
*
*
* *
*
*
*
0,00 50,00 100,00 150,00 200,00 250,00 300,00
1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 41 43 45 HS SCORE
KRAS
*
*
Heterogeneity Score (HS)
Normanno et al Ann Oncol 2015
Heterogeneity Score (HS) and efficacy of treatment in the CAPRI trial
Case 177 (SD, PFS 5,9 mo)
70% tumor cells
HS 14,29 KRAS G13D HS 17,14 PIK3CA ex20 HS 54,29 BRAF V600E HS FBXW7 R465H 48,6
Normal BRAF
KRAS PIK3CA
Case 118 (PR, PFS 3,9 mo)
70% tumor cells
HS 22,86 KRAS G12D HS 74,29 PIK3CA ex9 HS 57,14 TP53
Normal TP53
KRAS PIK3CA
FBXW7 Normanno et al Ann Oncol 2015
Challenges in genomics-driven oncology
• Alterations of “actionable” oncogenic pathways are not always predictive of response to targeted agents
• Different molecular alterations of the same oncogene may not be equivalent
• Intra-tumor heterogeneity may affect response to targeted agents
• The molecular profile of solid tumors may significantly
change following treatment with target based agents
Resistant cells are selected by
treatment with target based agents
Sensitive Resistant
Before Target Therapy
After Target
Therapy
Normanno J Cell Biochem 2013
The future of biomarker testing in cancer
A patient story in 2014
• A 39-year-old woman, was referred to S. Orsola Malpighi oncology unit in November 2013 for stage IV lung cancer.
• Total body CT scan showed: massive expansive process that occupies much of the middle lobe and basal portion; enlarged lymphnodes both on mediastinum and mesentere; neoformations of both adrenals;
presence of two nodular brain.
• 18 FDG-PET additionally showed intense metabolic activity of bone (D7, L4, 3 and 10 right rib), liver (segment 7) and peritoneum.
• She underwent to a needle biopsy of the lung with a diagnosis of undifferentiated carcinoma (immunohistochemistry negative for CD 117, synaptophysin, CD30, S100, calretinin, TTF1, smooth muscle actin;
partially positive for vimentine ed EMA; positive per citokeratin 8). A second histological revision suggested for a high grade sarcomatoid carcinoma.
A patient story in 2014
• Between November and December 2013 she underwent to a first line of chemotherapy with cisplatin (60 mg/mq), epirubicin (60 mg/mq) and ifosfamide (5000 mg/mq) but after only two cycles has been progressing on all sites of disease (fig.2).
• Subsequently she was subjected to a second line of treatment with
carboplatin (AUC 5) and paclitaxel (175 mg/mq). After three doses, there was a rapid clinical deterioration for worsening of intestinal
subocclusion due to carcinomatosis.
• Request of wide molecular screening at the Laboratory of Pharmacogenomic of CROM
BRAF mutations and response to BRAF inhibitors in non-small-cell lung cancer
Case DM5118
80% tumor cells HS 45 BRAF V600E
HS 1,25 p.E746_A750Del EGFR
Normal BRAF
EGFR Unknown
A patient story in 2014
• Off-label vemurafenib was started in March 2014 at a dose of 720 mg administered twice per day
• After 10 days of administration she had an admission to emergency room for an epileptic epidode with aphasia associated to hyperpyrexia (38°C). CT scan of the brain showed perilesional edema and she
underwent to whole brain radiotherapy
• After one month of treatment a total body CT scan showed: dramatic reduction of the process of the right middle lobe (8x6.7 cm vs 12x7.7 cm) and reduction in number and dimension of limphnodes and other
metastatic sites
• The patient suddenly died on May 2014 due to progression of cerebral lesions
Garraway JCO 2013
Surgeon Endoscopist
Radiologist
Surgeon Endoscopist
Radiologist
Medical Oncologist
Medical Oncologist
Pathologist, Molecular Biologist Medical
Oncologist
SurgeonRadiotherapist
