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Measuring Angiogenesis in Hematologic Diseases Maher Albitar, MD Quest Diagnostics, Nichols Institute San Juan Capistrano, CA 92690 (949)728-4784

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Presentation on theme: "Measuring Angiogenesis in Hematologic Diseases Maher Albitar, MD Quest Diagnostics, Nichols Institute San Juan Capistrano, CA 92690 (949)728-4784"— Presentation transcript:

1 Measuring Angiogenesis in Hematologic Diseases Maher Albitar, MD Quest Diagnostics, Nichols Institute San Juan Capistrano, CA (949) Steve Potts, Ph.D. Ravi Dayana Allen Olson, Ph.D. Aprio

2 Angiogenesis in Bone marrow

3 Variation in Angiogenesis Between Patients with Leukemias

4 1 CLL AML 6NN VEGF ACTIN CML-Cells CML-Plasma kD Expression of Vascular Endothelial Growth Factor (VEGF) in Leukemic Cells

5 Angiogenesis Vs. Hematpoiesis in Hematologic Diseases: Proangiogenic/Anti-angiogenic Factors: VEGF (multiple forms), bFGF, tal-1, Cox2, tie1, PIGF, PD-ECGF, HGF, MMP, PDGF, TNF-a, Angiogenin, TGF-a/b, G-CSF, VEGFR1/VEGFR2, NP1/NP2,..... Antiangiogenic Agents: Antibodies: Anti-VEGF, CDP860, Cotara, Erbitux.. Small Molecule Tyrosine Kinase inhibitor: AZD2171, BAy , PTK787/ZK, SU Other: Thalidomide,… Endothelial Cells Autocrine Paracrine Leukemia/Lymphoma Cells Stromal Cells

6 -IHC: FVIII antigen-positive endothelial cells -Digitized images are analyzed for: 1.Number of blood vessels/10 fields (200X) 2.Lumen area/10 fields (200X) using NIH Image software Increased Angiogenesis in Bone Marrow in Patients with Hematologic Diseases (Manual Measurement)

7 High Levels of Plasma Angiogenic Factors in Leukemias and MDS

8 Intracellular VEGF is an Independent Prognostic Factor in AML and MDS

9 Poor Reproducibility Time consuming and laborious Requires pathology background Not cost-effective Of marginal clinical Value No standardization How about Automation? Accuracy? Reproducibility? Clinically useful? Standardization between laboratories Measuring Angiogenesis (Microvessel Density)

10 Define sampling area: 5-10 rectangles representative of the sample (Automation in progress) Automation in Measuring Angiogenesis Calculate the space in these rectangles

11 Measuring Vascular Areas by Measuring Factor VIII+ Area Highlight brown (DAB)-stained Potentially: Define micro vessel boundary Find lumen regions (Holes) within micro vessels

12 Subtracting Megakaryocytes from Measured Vascular Area Hematoxyline stain with heavy Anti-Factor VIII are filtered out with colocalization algorithm

13 Find Micro Vessel Boundary Find Lumen regions Watershed segmentation Double Threshold Find & Fill Lumen Holes Find Micro Vessels (watershed) Measuring Partial Vascular Areas

14 Measuring Vascular Area relative to Hematopoietic Area/Cells (Vasc/Hem) Hematoxyline-stained area or individual cells can be counted in the same rectangles tested for angiogenesis.

15 High Concordance Between Hmatopoietic Cell Number and Hematopoietic Area

16 Significant Correlation Between Manual and Automated Vascular Area Measurements

17 Correlations of Automated Angiogenesis Measurements

18 Clinical Correlations in Patients with Acute Myeloid Leukemia and Myelodysplastic syndrome (MDS)

19 Distribution of Vascularity and Vasc/Hem in Patients with AML

20 Distribution of % Vascularity and Vasc/Hem in Patients with MDS

21 Higher Vasularity (% Vascular Area) is Associated with Longer Survival in MDS

22 Higher Vasular/Hematopoietic Area is Associated with Longer Survival in MDS

23 No correlation between Angiogenesis and Survival in AML

24 Higher Vasular/Hematopoietic Area is Associated with Shorter Remission Duration in AML

25 Summary Measuring Vascularity may Provide a surrogate marker for clinical behavior Automation in measuring angiogenesis is a reliable means for standardizing measurement of angiogenesis Automation allows the measurement of: Vascular Area (%) Hematopoietic cellularity Vascular Area relative to cellularity (Vasc/Hem) Larger studies are needed for clinical and biological correlations Further studies are needed to correlate with response the anti-angiogenic therapy


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