Medical Imaging Workshop Molecular Imaging Marcelo Tatit Sapienza INFIERI Summer School Intelligent signal processing for FrontIER Research and Industry 

Molecular Imaging Overview Imaging Modalities Clinical Applications – e.g. breast cancer

Molecular Imaging In vivo Imaging MOLECULAR BIOLOGY visualisation, characterization and quantification of normal / pathological biological processes at the cellular and molecular level Molecular Imaging emerged as a discipline at the intersection of molecular biology and in vivo imaging

 MOLECULAR BIOLOGY Molecular paradigm of diseases Abnormal cells with pathological phenotypes Molecular expression

Hallmarks of cancer – Cell 2000 Hanahan & Weinberg

Probes / ligands may be detected and allow Abnormal cells with pathological phenotypes Molecular expression Probes / ligands may be detected and allow Therapy with labeled compounds Diagnosis Identification of targets for drugs Therapy response Therapy planning

BASIC / PRECLINICAL RESEARCH CLINICAL APPLICATIONS Molecular Imaging BASIC / PRECLINICAL RESEARCH Study of mechanisms of disease development and progression Detection and activity of receptors and pathways Pharmacokinetics / pharmacodynamics of target drugs CLINICAL APPLICATIONS Understanding pathophysiological mechanisms Diagnosis / Staging Response to target drugs / individualized therapies

Translational research Preclinical Molecular Target Identification Development of ligands Experimental / preclinical evaluation Clinical Image in humans  validation Approval by regulatory agencies Clinical application

Translational research from BENCH to BEDSIDE to public health  In a medical research context, it aims to "translate" findings in fundamental research into medical practice and meaningful health outcomes

Molecular Imaging Overview Imaging Modalities Clinical Applications – e.g. breast cancer

Imaging Modalities Optical systems Nuclear Medicine: PET / SPECT MRI Ultrasonography Computed tomography Differences in Spatial resolution Depth of evaluation Ionizing / non-ionizing radiation Available molecular markers or probes Detection threshold

Imaging modalities Willmann Nature Reviews 2008

Imaging modalities Optical Imaging: lower cost  high-throughput screening for targets low depth penetration  limited clinical translation Nuclear Medicine: higher cost than optical unlimited depth penetration  clinical translation MRI: high resolution and soft tissue contrast / cost and imaging time US: high spatial and temporal resolution / low cost / limited targets CT: high spatial resolution / no target specific imaging Willmann Nature Reviews 2008

Spectrum of wavelenghts Eletromagnetic radiation MRI Optical CT / NM High energy Low energy Infra red Ultra violet

Optical Imaging fluorescence and bioluminescence Reporter gene Green fluorescent protein Near Infrared fluorphores (NIR) Reporter gene (luciferase) Prescher Current Opinion in Chemical Biology 2010

NM Radiopharmaceuticals radiolabeled molecules designed for in vivo application: PHARMACEUTICAL= molecular structure determining the fate of the compound within the organism RADIO= radioactive nuclide responsible for a signal detectable outside of the organism e.g. technetium-99m half life 6 hours gamma-ray photon 140 keV

Scintillation camara Sorenson and Phelps,27 1987 W.B.Saunders

SPECT Single Photon Emission Computed Tomography

Positron emitters Nuclides half life Positron: F-18 110 min C-11 20 min N-13 10 min O-15 1 2 min Ga-68 68 min Rb-82 1.3 min Positron: Same mass as electron opposite electrical charge annihilation generates a pair of gamma-ray photons – 180º

PET Zanzonico Semin Nucl Med 2004

SPECT PET 511 keV 140 keV SPECT / CT PET / CT

PET SPECT PET > SPECT Spatial resolution (human studies) Temporal resolution Sensitivity Cost

Molecular Imaging Requirements Imaging equipment Target selection Development of imaging probe / tracer

Development of in vivo probes < 5% of in vitro targets allow development of an in vivo tracer High TARGET concentration Affinity and specificity Absence of biological barriers (i.e. endothelium, blood brain barrier, ...) Stable labeling of compound

Development of in vivo probes < 5% of in vitro targets allow development of an in vivo tracer High TARGET activity / concentration Affinity and specificity Absence of biological barriers (i.e. endothelium, blood brain barrier, ...) Stable labeling of compound Low BACKGROUND activity Non-specific accumulation, Circulating or interstitial activity Renal or hepatic elimination

Development of in vivo probes < 5% of in vitro targets allow development of an in vivo tracer High TARGET activity / concentration Affinity and specificity Absence of biological barriers (i.e. endothelium, blood brain barrier, ...) Stable labeling of compound Low BACKGROUND activity Non-specific accumulation, Circulating or interstitial activity Renal or hepatic elimination Signal amplification Cell trapping Enzymatic conversion "Reporter" molecules: fluorescence, radiation, magnetic

18FDG fluorodeoxyglucose = glucose analogue EXAMPLE: 18FDG fluorodeoxyglucose = glucose analogue Transport (Glut) Phosphorylation (hexokinase) Metabolism

MOST TUMORS: Increased Aerobic glycolysis (Warburg effect ) Phenotype common to most tumors Lower production of energy / mol X NADPH Production - Synthesis Hypoxia and acidosis select cells resistant to apoptosis Acid pH associated with invasion Vander Heiden Understanding the Warburg Effect Science 2009

Hanahan & Weinberg Cell 2011 The same authors revisited the main molecular factors related to cancer, increased glycolisis is part of deregulated cellular energetics

Molecular Imaging Overview Imaging Modalities Clinical Applications – e.g. breast cancer

Breast cancer Brazil Most incident in women ~ 50 /100,000 57.120 new cases ( 2014 – INCA ) deaths: 13.345 ( 2011 – SIM ) 5 y survival ~ 60 % LOBULAR DUCTAL

Breast cancer Staging PROGNOSIS and CONDUCT - T 1 < 2 cm T2 2-5 cm T3 > 5 cm T4 thoracic wall / skin - N0, 1 axillary I-II mobile, N2 axillary fixed or int.thoracic, N3 infra (III) / supraclavicular / axillary+int. thoracic - Metastases M0, M1 AJCC Cancer Staging Manual. 7th ed. 2010, PROGNOSIS and CONDUCT Therapy choices considers also : Clinical conditions, Age , Menopause, Histology of the tumor Hormone Receptors and HER2

Hormone and Growth Factor Receptors expression variation PREDICTIVE biomarker = susceptibility of the tumor before indicating the therapy

personalized cancer therapy BIOPSY: TU hormone receptor ++  susceptible to treatment with drugs that blocks either the estrogen receptors or hormonal synthesis Biomarker-driven personalized cancer therapy Precision medicine  It is well known that gene expression may vary in different areas of the same tumor in a single patient. This publication from 2012 shows a primary renal cell tumor with differences in tumor grade from well differentiated to undifferentiated tumor – the metastases may also present marked heterogeneity. This heterogeneity results from the interaction of the tumor cells with its surrounding, acting in a Darwinian selection of clones, form which some are capable of dissemination and metastatic progpagation, others may be quiescent but resistant to a specific therapy, others may act as tumor stem cells. BUT…

Establishing genetic and molecular profile by biopsy may not be sufficient: Tumor heterogeneity Gerlinger, Intratumor heterogeneity NEJM 2012

 target = hormone receptor 18FES – FLUOROESTRADIOL  target = hormone receptor FDG FES FDG post-therapy PREDICTIVE biomarker in breast cancer ( indicates susceptibility to treatment ) Linden JCO 2006

18FES – FLUORO ESTRADIOL FDG FES FDG post-therapy Linden JCO 2006

PET- FDG in the metabolic evaluation after lymphoma chemotherapy EARLY RESPONSE biomarker = post-therapy prognosis PET- FDG in the metabolic evaluation after lymphoma chemotherapy Reduce or increase # chemotherapy cycles Change / add therapy Kasamon JNM 2007

18F-FES – FLUOROTHYMIDINE  target = DNA synthesis uptake after 1st cycle identifies responders ( p 0.001 ) - ( n= 15 ) EARLY RESPONSE biomarker in breast cancer Crippa F Eur J Nucl Med Mol Imaging 2015

18F-FES – FLUORO THYMIDINE EARLY RESPONSE biomarker in breast cancer uptake after 1st cycle identifies responders ( p 0.001 ) - ( n= 15 ) Crippa F Eur J Nucl Med Mol Imaging 2015

Conclusion Molecular imaging is a multidiciplinary field in the intersection of molecular biology and in vivo imaging Main pillars of MI are : Use of imaging modalities with different performances Development of probes/ligands detectable in vivo MI is part of translational research and may be applied for biomarker-driven personalized therapy ( precision medicine )

Thank you ! marcelo.sapienza@hc.fm.usp.br