Platelet Activation and Aggregation in Patients with Advanced Adenocarcinoma Undergoing Chemotherapy: Correlation with a Validated Venous Thromboembolism Risk Score

Background: Cancer is known to increase risk of venous thromboembolism (VTE), which is associated with considerable mortality and morbidity. Chemotherapy is an independent risk factor for thrombosis in cancer patients. We examined platelet activation and reactivity based on the Khorana Score, a validated scoring model for VTE risk in patients receiving chemotherapy.

Materials and Methods: Patients (n=25) with advanced stage adenocarcinomas (TNM stage III or greater), were enrolled in this study. Patients on antiplatelet therapy or on anticoagulation were excluded. Approximately 10 ml of Citrated blood samples were collected from central access catheters after discarding initial 5 ml to minimize iatrogenic platelet activation. 1μMol, 2μMol, 5μMol of ADP-induced, 2μMol Arachidonic Acid (AA)-induced  and 4μMol Collagen-induced platelet aggregation in Platelet Rich plasma (PRP) was assessed with a light transmission aggregometry assay. Maximal aggregation and aggregation velocity were recorded. Concurrent flowcytometry analysis was done to assess the expressions of CD41 (Glycoprotein GpIIb), CD62p (platelet surface p-Selectin), and PAC-1(Activated GP IIb/IIIa). 

Thromboembolic risk scores (Khorana Scores, KS) were calculated based on a validated scoring system (Khorana AA, et al. Blood. 2008;111:4902-7) as follows:

Very high risk cancers (pancreatic or gastric)	+2 points
High risk cancers (lung, ovarian, or bladder cancer)	+1 point
Platelet count ≥350 x 109/L	+1 point
Hemoglobin < 10 gm/dL or use of erythropoietin	+1 point
Leukocytosis >11 x 109/L	+1 point
BMI  > 35	+1 point

Results: Mean values for maximum platelet aggregation were calculated by groups and compared between patient with KS < 3 and KS > = 3 by two sample t-tests. Differences were statistically significant for all concentrations of ADP and collagen, with a positive trend for Arachidonic acid (Table). A significant linear relationship between maximal platelet aggregation and higher KS was observed. However there were no significant differences observed in the expression of platelet surface p-selectin, CD41 or PAC-1 when comparing patients at Khorana scores 0, 3 or more than 3 (not shown).

Agonist Concentration	Khorana Score <3	Khorana Score >= 3	P value
ADP 1μMol	20.8%	28.14%	0.02
ADP 2μMol	38.6%	67%	0.002
ADP 5μMol	66.5%	79.85%	0.006
Arachidonic acid 2μMol	61.6%	73.85%	0.12
Collagen 4μMol	78.6%	90.14%	0.01

Discussion: The mechanism of cancer-related thromboembolism is not well understood. Chemotherapy is an added risk factor for the development of VTEs. Several new markers such as soluble p-selectin and mean platelet volume have been investigated as adjunct factors to improve predictive ability. Although our study did not find a correlation of some common platelet surface markers for activation with thrombotic risk scores; we find a strong positive correlation of future thrombotic risk derived via Khorana scores with heightened platelet reactivity. Platelet reactivity may mediate a final common pathway for venous thromboembolism in cancer patients and should be validated further as an adjunctive marker in a large scale study.