SPARC in Pancreatic Cancer
• utilises the properties of albumin to reversibly bind paclitaxel, and transport it across the endothelial cell and concentrate it in areas of tumor
• avoids the use of Cremophor EL, which contributes to serious toxicity (e.g. hypersensitivity, axonal degeneration) and requires special infusion tubing, premedication and prolonged infusion
• shows improved bioavailability and linear pharmacokinetics, whereas CrEL forms micelles entrapping the paclitaxel, leading to decreased unbound drug fraction, decreased drug clearance and lack of dose-dependent antitumour activity
Nab-paclitaxel properties
Solvent-based taxanes provoke formation of micelles in circulation
• Micelle formation in the circulation entraps paclitaxel in plasma
• Resulting non-linear pharmacokinetics contribute to a lack of dose-dependent antitumour activity
Aapro et al. EJC Suppl. 2008;6:3–11 Hamad et al. Expert Opin Drug Deliv. 2008;5: 205–219
Largemicelle
Control plasma Plasma + solvent-based paclitaxel
• The accumulation of paclitaxel in tumors was 33% higher for nab-paclitaxel compared with paclitaxel (P < .0001)1
nabnab -Paclitaxel Results in Higher Tumoral Uptake -Paclitaxel Results in Higher Tumoral Uptake Compared With paclitaxelCompared With paclitaxel
Desai et al. Clin Cancer Res. 2006; 12:1317-1324.
Hours
Pac
litax
el (
nC
i/g)
140
80
60
400.1 1 10 1000.01
120
100
Tumor AUC nab-paclitaxel =1.33 x Taxol
p < .0001 ANOVA
nab-paclitaxel
Taxol
AUC(nCi•hr/g)
3,632
2,739
Ka(hr1)
0.43
0.13
AUC, area under the curve; KA, absorption rate.
Tumor Uptake in Nude Mice Xenografts Following 20 mg/kg Dose of Paclitaxel
KEY STEPS in albumin-paclitaxel delivery to tumor
• Albumin initiates the endothelial transcytosis of paclitaxel by binding to a cell surface receptor: 60-kDa glycoprotein (gp60).
• In turn, gp60 associates with caveolin-1 resulting in the invagination of the endothelial cell membrane trapping the complex in vesicular structures called caveolae.
• Clustering of the gp60-albumin complex during vescicle formation reduces receptor affinity for albumin, which permits the release of albumin and any bound ligands to the abluminal side of the cell.
• Albumin accumulates in tumors, possibly due, in part, to the secretion of the albumin-binding protein SPARC (secreted protein, acidic and rich in cysteine, osteonectin or BM-40), which, in turn, may result in preferential intatumoural accumulation of albumin-bound molecules.
The Unique Properties of Albumin Improve the Risk to Benefit Profile of nab-Paclitaxel
• 130-nm sized albumin-paclitaxel complexes1,2
– nab-Paclitaxel is the first nanotechnology-derived agent approved for the treatment of breast cancer
– Albumin gives nab-paclitaxel linear pharmacokinetics3 = predictable drug exposure with dose modification
1. Desai et al. SABCS. 2004 [abstract 1071].2. Kratz et al. J Control Release. 2008;132(3):171-183.
3. Ibrahim et al. Clin Cancer Res. 2002;8(5):1038-1044.nab® is a registered trademark of Celgene Corporation.
Albumin
Paclitaxel
nab-Paclitaxel particle
2D Conceptualization
Mechanism of Action of nab-Paclitaxel
Investigation of the functional importance of SPARC with respect to nab-paclitaxel is ongoing.
SPARC, secreted protein acidic and rich in cysteine.
Albumin-Mediated Transcytosis of Paclitaxel
SPARC, Secreted Protein Acidic and Rich in Cysteine.
Endothelial cells
Tumor cells
Subendothelial space
Investigation of the functional importance of SPARC with respect to nab-paclitaxel is ongoing.
MDA-MB-231
0 10 20 30 400
500
1000
1500
2000
2500
3000
SalineAbraxane 15 mg/kg q4dx3
Days
Tu
mo
r V
olu
me (
mm
3)
Low Medium HighSPARC SPARC SPARC(TGI = 36%) (TGI = 60%) (TGI = 81%)
SPARC level in heterogeneous tumors affects SPARC level in heterogeneous tumors affects relative response to relative response to nabnab-paclitaxel-paclitaxel
• Stromal SPARC was associated with worse outcome and poor survival
• Tumoral SPARC did not correlate with survival
Peritumoral Fibroblast SPARC Expression and Patient Outcome With Resectable Pancreatic Adenocarcinoma
Infante 2007
Overexpression of SPARC gene in human gastric carcinoma and its clinic-pathologic significance
Non-cancerous Mucosa Gastric Cancer
Diffuse Type
Intestinal Type
Wang, C-S et al - British Journal of Cancer (2004) 91, 1924 – 1930
Summary of SPARC as a marker of poor prognosis
Classification SPARC Expression/ Function Reference
Hepatocellular Carcinoma
Overexpression by stromal myofibroblasts correlates well with angiogenesis & tumor
progression Lau et al. 2006
Glioblastoma Overexpression in juxtratumoral perivascular cells but not non-malignant brain vessels
Pen et al. 2007
Multiplle Myeloma Significant decrease in plasma levels of SPARC has a prognostic value & shows + correlation
with Hb levels & platelet counts Turk et al. 2005
Meningioma A diagnostic marker for invasive meningiomas regardless of grade
Remple et al. 1999
Prostate Carcinoma High levels of SPARC mRNA & protein as a marker of CaP metastatic foci
Thomas et al. 2000
Head & Neck Cancer High/ Marker of poor prognosis Chin et al. 2005
Tongue Carcinoma High/ Marker of poor prognosis Kato et al. 2005
Cervical Carcinoma High/ Marker of poor prognosis Sova et al. 2006
Non-small cell lung cancer
High/ Marker of poor prognosis Koukourakis et al. 2003
Bladder Cancer High/ Marker of poor prognosis Yamanaka et al.
2001
Melanoma High levels correlate with metastasis Massi et al.
1999
Esophageal Cancer High/ Marker of poor prognosis Yamashita et al.
2003
Breast Cancer High/ Marker of poor prognosis; shows + correlation with stage & grade
Watkins et al. 2005
SPARC Expression in Microarrays Performed on SPARC Expression in Microarrays Performed on Tumors Taken Directly From Patients Tumors Taken Directly From Patients
SPARC Expression in Microarrays Performed on SPARC Expression in Microarrays Performed on Tumors Taken Directly From Patients Tumors Taken Directly From Patients
Tumor Type # with SPARC /# studied (%)
Breast 6/9 67%
Ovary 5/15 33%
Pancreas 13/16 81%
Melanoma 15/17 88%
Adrenal 2/5 40%
Colon 4/15 27%
Total† 76/113 67%
*Increased expression at level of 0.001 versus normal tissue
Slide courtesy Dan Von Hoff, AACR 2006 Slide courtesy Dan Von Hoff, AACR 2006
Preclinical Platform
Rubio et al, CCR 2006
0
10
20
30
40
50
60
GEM ABI GEM+ABI
Tu
mo
rs r
egre
ss
ed
50
% o
f it
s in
itia
l siz
e (
%)
Average Response Rate in Xenografts (n = 11)Average Response Rate in Xenografts (n = 11)
1. Pancreatic cancers are poorly perfused
2. One factor – an intense fibro inflammatory reaction – stroma – squeezes out blood supply and stops infiltration of immunocytes
3. Need to attack the stroma to improve tumor cells killing
• have a poor blood supply
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On a Microscopic levelOn a Microscopic level
Effects of Effects of nabnab-paclitaxel on Tumor Stroma-paclitaxel on Tumor Stroma
Collagen Type I Staining
B
Effects of Effects of nabnab-paclitaxel on Blood Vessels-paclitaxel on Blood Vessels
0
1000
2000
3000
4000
5000
6000
7000
8000
GEM
Con
cent
ratio
n ng
/g tu
mor
(Mea
n ±
SE
M)
GEM alone
GEM+ABI
Effects of Effects of nabnab-paclitaxel on Gemcitabine Delivery-paclitaxel on Gemcitabine Delivery
• Role and regulation of SPARC in Pancreatic Cancer.
• SPARC as • prognostic (which patients need rx) • predictive biomarker (which patients are likely to benefit
from a specific rx).
• Stromal effects.
• Diagnostic test.
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Questions in DevelopmentQuestions in Development
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Hypothesis: Albumin-Binding Proteins May Aid in the Uptake of
nab-paclitaxel Into Tumors
• Accumulation of albumin in tumors may be mediated by the protein SPARC1
– SPARC binds albumin2
– Many tumor types overexpress SPARC compared with normal tissues3-5
• High SPARC expression has been shown to be a negative prognostic indicator in many cancer types4,5
1. Kratz F, et al. J Controlled Release. 2008; 132:171-183.2. Schnitzer JE, et al. J Biol Chem. 1994;269(8):6072-6082.
3. Watkins G, et al. Prostaglandins, Leukotrienes and Essential Fatty Acids. 2005;72:267-272.4. Desai N, et al. Transl Oncol. 2009;2:59-64.
5. Podhajcer OL et al. Cancer Metastasis Rev. 2008;27:691-705. SPARC, Secreted Protein Acidic and Rich in Cysteine.
SPARC Expression and Clinical ResponseD. von Hoff et al. ASCO 2009 Poster # 4525
SPARC status by IHC was available for 32 RECIST evaluable patients (investigator dataset: 2CR, 14PR, 14SD, 2PD)
Staining of tumor cells (and not stromal fibroblasts) by antibody P showed improved response for SPARC+ patients (P = 0.027)
Other epitopes of SPARC showed similar response between SPARC+ve and SPARC-ve groups (P = NS)
Fraction (%) of Patients Responding
SPARC status Antibody P (Tumor Cell)
(N=32)
Antibody M (Tumor Cell)
(N=32)
Antibody P (Stromal Fibroblasts)
(N=27)
Antibody M (Stromal Fibroblasts)
(N=27)
SPARC Positive 8/10 (80%) 2 CR / 6 PR 2/5 (40%) 8/16 (50%) 3/5 (60%)
SPARC Negative 8/22 (36.4%) 0 CR / 8 PR 14/27 (52%) 7/11 (64%) 12/22 (55%)
P-value 0.027 NS NS NS
P positive pt # 014 P negative pt # 012Abbreviations: M = antibody M; NS = not significant; P = antibody P
1. Celgene data on file.
Though high SPARC expression is typically a poor prognostic factor, it actually predicts an improved response to nab-paclitaxel in this trial in terms of overall survival
SPARC status was evaluated in 36 patients:a significant increase in OS was observed for patients in the high-SPARC group versus the low-SPARC group (median OS: 17.8 vs 8.1 months, respectively, P=0.0431)
SPARC Expression and Clinical ResponseD. von Hoff et al. JCO 2011
SPARC Expression and Clinical ResponseD. von Hoff et al. JCO 2011
SPARC Assessment: actual tools• Several antibodies are available and have been
used in publications assessing SPARC protein• Antibodies have different staining patterns, thus
need to optimize an assay:– Compare available antibodies– Standardize antigen retrieval, antibody dilution,
secondary antibody and automated staining systems.– Scoring criteria needs to be simplified and
standardized– Tumor compartments staining with SPARC will vary
based on histology
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Biomarker Development: Requirements
Generation of a biomarker hypothesis
Develop prototype assay
Identification of candidate markers
Discovery
Clinical validation of candidate m. using assay
Assay refinement & development IVD test
Regulatory test approvalValidation
Distribution to laboratories
Auditing for result consistency
CommercialisationClinician/Lab education
Biomarker evaluation in CA046
• Archival tumor tissue for SPARC IHC– Slides and/or blocks for received from 160
patients to date– Further explore role of SPARC in response to nab-
paclitaxel in context of a randomized trial
27
Next steps in developing SPARC prototype assay
• The prototype assay is based on IHC methodologies– a collaboration with EU and USA academic
centers has been established with the goal to define the IHC platform
– M. Hidalgo, Madrid, will lead this collaboration, and the kick-off meeting will take place October 17 in Seville
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SPARC: where we are today
Potentially valuable
Early stage assessment
Clinical use
Exploratory: current standing of SPARC assessment
In Summary
• Stromal components are strategic targets in Pancreatic Cancer.
• SPARC is a stromal component.
• Gem-Nab-Paclitaxel promising activity.
• Stromal depletion vs stroma modification.
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Back-up
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SPARC IHC method• Developed at MPI (AZ); transferred to
St.John’s pathology lab (CA)• Two antibodies used; each slide scored
separately– Monoclonal R&D #MAB941(1:250; 30 min)– Monoclonal Haematologica Technologies, Inc,
#A0N5031 (1:250; 30 min)– Polyclonal also evaluated, but not used
• R&D AF941 (1:150; 30 min)
• DAKO Automated stainer, NO Antigen retrieval
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Scoring criteria
• Tumor compartments assessed separately– Tumor, fibroblasts, blood vessels, inflammatory
cells, background stroma tissue, any normal tissue
• Highest intensity (0 to +4) for a compartment (>10% of cells in that compartment) recorded
• % staining at highest intensity in compartment recorded
33
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