world ctc berlin 2013
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CSFTCs can be detected, quantified andTRANSCRIPT
Detection and quantification of Tumoral Cells in CSF (CSFTCs): E.Le Rhun (Lille) GC Faure (Nancy)Nancytomique CHU Nancy
Diagnosis of leptomeningeal metastases (LM) in patients with solid tumors (breast, lung, ...) and melanomas remains difficult.
Usual diagnostic methods of cytomorphological assessment of cerebro-spinal fluid (CSF) and gadolinium enhanced MRI lack both specificity and sensitivity.
After CTCs.......... CSFTCs
• A new acronym– CSF:volume 150 mL vs blood (4,5L)– Another biological fluid– From choroid plexuses to pathology
• A new gold standard for carcinoma meningitis or leptomeningeal metastasis definition
• A new frontier for cancer research– Metastatic processus– New therapeutic approaches?
Cerebrospinal fluid: CSF
• Volume 150mL• Production #500mL per day, (3.7x)• Choroid plexuses
• Lumbar puncture Berlin– Heinrich Ireneus Quincke – Berl klin Wochenschr 1891;28:929 +965
Leptomeningeal Metastasis
• Clinic– Very sick: seizures, severe headaches,
blurry vision, mental status changes, inability to walk or perform everyday tasks... completely incapacitated
• Diagnosis– Imaging (MRI)
• Meningeal enhancement
– Cytology– Biomarkers? Molecular, Cellular
LM in Media, and Internet 3/2013
• Huffington Post• People: Valerie HarperHarper, famous as the spunky best friend
Rhoda Morgenstern on The Mary Tyler Moore Show, told People she was stunned after receiving her diagnosis, but realized she could help spread awareness for the rare condition. « I think there's an opportunity to help people! »
CSFTCs: a new frontier in Cancer?
- L. Nayak, M. Fleisher, R. Gonzalez-Espinoza et al. (MSK, NY) Immunomagnetic platform technology (IMPT) for the diagnosis of leptomeningeal metastasis in solid tumors (LMST) 2010 ASCO Poster Discussion Session, Abstract Number: 2032.
Neurology 2013, small heterogeneous series of 15 LM
- Patel et al Hershey (Oncotarget 2011 Oct;2(10):752-60)
Spiking in normal blood
- Burns TF, Wolff AC (Johns Hopkins, Baltimore) Cell Cycle. 2012 Jan 15;11(2):203-4. Epub 2012 Jan 15. Detection of circulating tumor cells in the cerebrospinal fluid: a new frontier.
LM: Epidemiology... Prognosis 3 to 5% of cancer patients, incidence up to 9.6% (J Clin
Oncol 2004;22:2865) Up to 19% of autopsied patients with cancer and
neurological symtoms (Glass, 1979)– Breast cancer (5%), lung (11%), melanoma (20%)
Increasing incidence – Better survival of cancer patients
+ New molecules for systemic disease have bad meningeal diffusion Kodack DP et al. PNAS 2012, 109, E3119
Very Bad prognosis (4 weeks to 6 months) and bad quality of life
– But promise of new intrathecal drugs (MTX, trastuzumab...) and trials (Chamberlain)
Epidemiology of LM
• Probably underestimated, but• 100 000 to 170 000 patients with cancer each year
in the USA develop CNS metastases (Clin Canc Res 2007, 13; 1648) (J Clin Oncol 2004;22:2865)
– Breast (5 %)– Prostate (less)– Lung (20%)– Melanoma (7%)– Renal (6%)– Colorectal (2%)
• With major quality of life consequences
Quality of life
• Literature is poor• Support Care Cancer 2011;19: 467-
473 (Lung cancer)
• Complaints: pain, fatigue, dyspnea• Symptoms
– related to brain tumors: consciousness deterioration, headache, cranial nerve palsy, delirium
– Carcinomatous meningitis: headache, cranial nerve palsy, epilepsy, nausea + vomiting
LM: Gold standardDux et al, J Neurol Sci, 1994; 121; 74-78
CSF volume– 3.5mL: 68% positivity – 10.5mL: 97% positivity
Time interval between sampling and analysis Cell viability 30 mns 50%; 60 mns 20%; 90 mns 10%
Good sensitivity requires First LP 40% Second LP 80%, Third LP to reach 90-95%
No reliable quantification– Response at 50% threshold
Methods
• >80 Samples from Lille and Nancy– Volume 5mL– Up to 4 days delay between sampling
and study...– Lumbar and ventricular punctures
• Cytology (on 7.5 to 10mL) and biochemistry
according to classical diagnostic procedures • Cellsearch® technology (CTCs kit;
CMCs kit)
Patients• Established or suspected LM from
primitive cancers– Breast (45), Lung (15), Prostate and Lung (1),
Melanoma (5), Ovary (1)– Cytologically defined (50%) and MRI+– Patients included in DEPOSEIN clinical
research protocol (14)– Patients sampled for diagnostic and follow-up
procedures, at time of intrathecal treatment
• Control patients sampled in context of other neurological disorders
CSFTCS Breast (BMC Clinical Pathology)
CSFTCs Lung
STA Ch 2: LCR mélanome CMCNew DevelopmentsCSF vs BLOOD
• Preservation: CSF paradox in Cell Save tubes!
• Morphology, numbers and cell biology characteristics
– Similarities: Breast– Discrepancies: Lung (+CTMs), Melanoma
Main Results Specificity: no contaminating ependymal cells
in controls Sensitivity: Detection and quantification in all
established LM patients studied– Initial point of follow-up
• From 1 to >10000 cells– Sequential study in 9 patients from
Deposein with #30 assays High homogeneity (and reproducibility) of
images in patients according to primitive cancer types
High purity compared to blood samples Presence of CTM in lung cancer
CSFTCs and CSFMCs numbers With the CellSearch® Veridex
Cancer type
Breast Lung Melanoma
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Sequential analysis of CSFTCs confirms repetability of numerations with two subgroups (high > 700/mL vs low)BMC Clin Pathol 2012
Sample number
1st 2nd 3rd 4th 5th
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/ 5 m
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DM CTCWA CTCCJ CTCDMB CTCVT CTCPV CTCBE CTCCS CTCHE CMCST CMCDC CTC
Melanoma Leptomeningeal metastasis: current status
• CMCs are not easy to detect in bloodwith Cell Search technology
• Meningitis is underdiagnosed with severe prognosis
– L Harstad et al: Neuro Oncol 2008; 10: 1010-8 MD Anderson
Melanoma CSFMCsMedical Oncology 2013;
• CMC kit (J&J, VERIDEX)• Four patients 9 points• Good reproducibility during follow-up• Cell morphology of melanoma cells in CSF far
better than in blood
LUNG CancerCSFTCs, CSFTMs
• Patients 15, samples 18• Numerous CSFTCs
– Sequential follow-up (3)– Cell galleries allow to differentiate
SCLC (1), NSCLC adenocarcinoma (4+), NSCLC squamous carc. (3)....
– Aspects of apoptosis, autophagy...
• Numerous CTMs in some NSCLC patients up to 80%
LUNG Carcinoma meningitis
• CSFTCs --->
• <---CSFTMs
CSFTCs: a new frontier!
• Tumoral (epithelial) cells can be detected and quantified in CSF with the CellSearch® technology (CSFTCs)
• Their numbers can be sequentially followed-up in breast, lung and other cancers
– allowing to evaluate the efficacy of treatments (intrathecal and/or systemic)
• Tumoral cell population in CSF might be different from blood CTCs, allowing further studies of metastatic properties
• CSFMCs can also be detected and quantified in CSF
Research (1)Clinical
• Validation of sensitivity and specificity CSFTCs and CSFMCs is underway
– Sensitivity and reliability of the method for detection of rare events invites to use it earlier in clinical evolution of metastatic cancers to detect infraclinic LM
– CTM-like in the CSF are detectable and can be quantified with the CellSearch technology. Are they prognostically significant?
Research (2)Understanding cancer biology
Tumour Cell characteristicsHER-2, EGF-R, etc
Fi Melanoma CTCs are expressing HER-2
• Are CTM-like agregates in the CSF the metastatic ones?
Nature Cell Biology 15 [3] (février 3): 317 324. doi:10.1038/ncb2681.
Research (3)Metastases through the BBB
How do cells migrate preferentially to the brain and leptomeninges?
• Breast: Dario MarchettiIn epithelial cell adhesion molecule
(EpCAM)–negative CTCs, ... identified a potential signature of brain metastasis comprising “brain metastasis selected markers (BMSMs)” HER2+/EGFR+/HPSE+/Notch1+
Others?
Research (4) drug screening... for new therapy targets
• In clinically established LM, CSFTCs are not rare events, and
– Cells available from CSF of LM patients for further studies will help detecting new molecules for systemic or local treatment
– Sensitivity and reliability of the method for detection of rare events invites to use it earlier in clinical evolution of metastatic cancers (breast, lung, melanoma...)
• Not only to detect infraclinic LM in order
• ...to try local treatments as soon as possible and evaluate their efficacy
Participants:
• Centre Oscar Lambret (Lille)• CHU (Nancy)
– Pôle Laboratoires (Immunologie)– Pôle Neurologie (L Taillandier, Internes:
Marie, Maud?, Basile...)• Université Lorraine: SIGRETO (F Plenat), CRAN
CNRS UMR 7039 (D Wolff)• Hôpital Zhongnan (ZHOU Yunfeng, TU
Jiancheng, XIONG Bin Wuhan University• NENO Network
– Amiens, Besançon, Colmar, Reims, Strasbourg... Luxembourg, Liège...
AcknowledgmentsEA 4369 RHEM UMR CNRS 7039
• GC Faure• M de Carvalho• MC Béné (Nantes)• Wuhan PhD students (Chen Min, Cai Huili, Tu Qian)
• Laboratoire d'Immunologie, CHU Nancy, Pôle Laboratoires et Faculté de Médecine, Université Lorraine