Is Ashwell-Morell Receptor Mediated Hepatic Clearance of Platelet Clinically Relevant in Immune Thrombocytopenia (ITP)? a Retrospective Analysis of Platelet Kinetic Studies and Autoantibody Type Data

Introduction. Fc-mediated clearance of platelet (plt) by splenic macrophages is a characteristic feature of immune thrombocytopenia (ITP) and has so far been addressed as the main mechanism of plt destruction. Recently (Li J et al. ASH 2014 abs#467), an Fc-independent mechanism has been proposed which involves mainly the liver, mostly mediated by anti-GPIb antibodies (Abs). These Abs were shown to induce plt activation leading to GPIb desialylation. In turn, plt lacking sialic acid are removed by the hepatic Ashwell-Morell receptor (AMR). In order to test the clinical relevance of this novel mechanism of plt clearance, plt kinetic studies (PKS) and Ab type data were retrospectively reviewed in adult ITP patients (pts).

Patients. Charts of ITP pts who had data on both PKS and Ab testing were retrospectively reviewed. Pts are enrolled in a local Italian data base and informed consent to clinical data use is given at enrollment.

Results. A total of 82 pts (M 34) were identified; median age at ITP diagnosis was 46.3 yrs (range 7.5-78.9 yrs); median disease duration at PKS was 12.5 mos (range 0.7-278.9 mos). Of the 82 pts included: 59 (71.9%) had already undergone splenectomy between year 2000 and 2015; 23 were studied but splenectomy has not been performed yet. All pts underwent or are candidate to splenectomy because of steroid-dependent ITP; 1 pt only was primarily refractory to steroids and IVIG treatment. Results of PKS and autoAb testing are summarized in Figure 1. Antibody testing: 45/82 (55.9%) of pts tested positive to one or more than one antiGP Abs (direct test); 10/82 pts tested positive to both direct and indirect test (not shown in Fig). PKS results: Ab negative pts: spleen 25/37, liver 0; mixed 12/37; Ab positive pts: spleen 25/45, liver 4/45, mixed 16/45.  A trend toward an association between number of Ab positivity and site of clearance was observed: every single unit increase in the number of Ab positivity (i.e.: none vs 1 vs 2 vs 3 vs both direct and indirect Ab testing) is associated with a 33% increase in the odds of liver involvement (WaldÕs test after logistic regression p= 0.104). No association was found between either isolated or combined anti-GPIb-lX Ab positivity and site of plt clearance (FisherÕs exact test p=0.235). Splenectomy outcome: CR 50/59; R 4/59 ; NR 5/59. (Fig. 2)

Conclusions. Recent studies have proposed that Ab type may determine pltÕs fate in ITP by activating either Fc- or non Fc-mediated mechanisms of plt clearance resulting in splenic or hepatic uptake respectively. However, our data on PKS and Ab type do not support such a mechanism being clinically relevant in ITP pts. Although an increase in the number of Ab positivity increases the probability of liver involvement in plt clearance, no association was found between  type/number (single vs multiple antiGP) of Ab positive test and site of plt clearance. Moreover, response to splenectomy doesnÕt seem to be influenced by either Ab type and/or site of plt clearance. However, it may well be that AMR-mediated hepatic plt clearance rather represents a physiological mechanism involved in plt and hemostasis homeostasis. Plt desialylate as they circulate, thereby becoming the primary AMR ligand and this interaction is involved in the regulation of TPO production by hepatocytes (Grozovsky R et al. ASH 2014, abs #2). Desialylation also occurs when plt are activated by several physiological stimuli (S¿rensen AL et al. Curr Opin Hematol, 2008) and AMR clearance of activated plt may be relevant in attenuating the coagulopathy associated with sepsis (Ellies LG et al. Proc Natl Acad Sci USA 2002; Grewal PK et al. Nat Medicine 2008)-

Fig 1
Ab pattern	n(%)		site of plt clearance
			(111-Indium labeled plt)
		splenic	hepatic	mixed
All patients	82 (M34/F48 )
Ab negative	37 (45.1)	25 (67.6%)	0 (0%)	12 (32.4.%)
Ab positive	45 (54.9)	25 (55.5)	4 (9%)	16 (35.5%)
one positive Ab	17/45
GpIIb-IIIa	11	7	2	2
GpIb-IX	4	2	1	1
GpIa-IIa	2	1	0	1
two positive Abs	9/45
GpIIb-IIIa & GpIb-IX	2	1	0	1
GpIIb-IIIa & GpIa-IIa	7	5	0	2
three positive Abs	16/38
GpIIb-IIIa & GpIb-IX & GpIa-IIa	19	9	1	9

Fig 2
Ab pattern	Outcome after	site of plt clearance(111-indium labeled plt)
	splenectomy
		spleen	liver	mixed
Ab negative (27/59)
	CR + R	17	0	9
	NR	1
one positive Ab (13/59)
IIb-IIIa*	CR	4	2	1
	NR	0	0	0
Ib-IX
	CR	2	0	1
	NR	0	1	0
Ia-IIa
	CR	1	0	0
	NR	0	0	1
two positive Abs (8/59)
IIb-IIIa; Ib-IX
	CR	1	0	1
	NR	0	0	0
IIb-IIIa; Ia-IIa
	CR+R	4	0	2
	NR	0	0	0
three positive Abs (11/59)
IIb-IIIa; Ia-IIa; Ib-IX
	CR	6	0	3
	NR	1	1	0
*4 pts had both direct and indirect positive ab testing