Changes in D-dimer levels were inversely correlated with changes in platelet counts
The early decrease in platelet numbers was neither associated with levels of D-dimer at study entry, nor with change in the hemoglobin levels [correlation = 0.02 (P = 0.66) and −0.06 (P = 0.06), respectively]. However, for drug conservation patients with D-dimer values available at baseline and either month 1 or month 2 (n = 75), the rank correlation coefficient between change in D-dimer and change in platelets was −0.24 (P = 0.04). The association was even stronger [rank correlation coefficient −0.41 (P = 0.02)] for drug conservation patients who were on ART and had a suppressed viral load at study entry (n = 33) (Fig. 2a). For drug conservation patients, the correlations between change in platelets and changes in IL-6 and hsCRP were −0.23 (P = 0.05) and −0.06 (P = 0.59), respectively. When restricting to those on ART with a suppressed viral load, the correlations are −0.29 (P = 0.11) for IL-6 and −0.01 (P = 0.97) for hsCRP.
Changes in HIV-1 RNA levels inversely correlated with changes in platelet count in drug conservation patients with initially suppressed viral loads
When restricted to drug conservation participants with HIV-1 RNA 400c/ml or less on ART at entry (n = 738), changes in HIV-1 RNA levels after interruption of ART were inversely correlated with changes in platelet count (Fig. 2b). The correlation between 4-month change in log10 transformed HIV-1 RNA and 4-month change in log10 transformed platelet count was r = −0.34 (P < 0.001).
Recovery of platelet counts following (re)initiation of antiretroviral therapy was dependent on size of prior decline in platelet count as well as HIV-1 RNA levels at time of (re)initiation
Of 900 patients in the drug conservation arm (re)initiating ART, 663 had platelet information available thereafter and had a study entry platelet level greater than the value prior to initiation. For these 633 participants, the median (IQR) change from study entry platelet level to level prior to initiation was −47 000 (−79 000, −27 000) and 489 (74%) recovered to their study entry levels during follow-up. Estimated percents recovered by 4, 8 and 12 months after ART reinitiation were 45, 60 and 69%, respectively. Those with the larger declines in platelet counts from study entry to ART initiation [aHR per log10 higher = 184 (56–610)] and those with higher HIV-1 RNA levels at time of (re)initiation [aHR per log10 higher = 1.3 (1.2–1.5)] recovered their platelet counts faster. Those who initiated ART later in the study took longer to recover their platelet counts (Table 3).
Thrombocytopenia and risk of subsequent clinical disease
In the drug conservation arm 199 of the 1010 participants with month 4 platelet levels (20%) had a platelet decline of at least 25% during the first 4 months. A 25% decline in platelet level at 4 months was not associated with subsequent risk of serious non-AIDS events [aHR = 0.8 (0.4–1.7), P = 0.52]. Developing thrombocytopenia was not associated with subsequent risk of serious non-AIDS events [aHR = 0.9 (0.6–2.6); P = 0.92].
Here we describe platelet kinetics, and the relationship to HIV replication and changes in D-dimer levels, in the context of an ART interruption study (SMART). A significant decrease in the platelet count as well as an increased rate of thrombocytopenia (defined as <100.000/μl) was observed among 1090 participants randomized to ART interruption (drug conservation arm) in the SMART Study, whereas patients assigned to continuous ART use (viral suppression arm) had stable platelet counts. Inverse correlations were seen between changes in platelets and changes in D-dimer or HIV-1 RNA levels, and were strongest among those with viral suppression at study entry who then stopped ART. Most patients recovered to platelet levels seen at study entry after reinitiating ART.
The prevalence of HIV-1-induced thrombocytopenia varies greatly between different studies, with reports between 1.1 and 54.7%, but is generally more than 10% . HIV-related thrombocytopenia is likely caused by accelerated platelet destruction, splenic platelet sequestration, and variably impaired platelet production . Accelerated clearance can be caused by immune-complex-based disease, antiplatelet glycoprotein antibodies, and/or anti-HIV-1 antibodies that cross-react with platelet membrane glycoproteins . Impaired platelet production is probably caused by HIV-1 directly infecting megakaryocytes through the viral CXCR4 coreceptor, inhibiting mature megakaryocytes from effectively producing platelets . HIV-1 can also impair megakaryocyte maturation as shown by reduced formation megakaryocytopoietic colony forming units in HIV-1-infected individuals .
The predominant mechanism(s) contributing to HIV-related platelet reductions may differ by HIV disease stage. In the early phase of disease with low viral loads, an immune thrombocytopenic purpura (ITP) like mechanism with presence of antiplatelet antibodies and increased platelet destruction dominates. In advanced HIV-1 infection with high viral loads and low CD4 cell counts (<200/μl) thrombocytopenia is mainly caused by decreased platelet production [15,16].
HIV-1-induced thrombocytopenia is usually responsive to standard ITP treatments (prednisone, intravenous immunoglobulin, anti RhD, splenectomy). Moreover, reducing viral load using ART has also proven efficient [12,17].
Our study is one of the few available wherein incidence and risk factors of incident thrombocytopenia have been systematically compared in the context of continuing/starting and stopping ART within a randomized trial. The findings are consistent with the Agence nationale de recherches sur le sida en les hépatitis virales (ANRS) 106-window trial analyzing 391 patients randomized to continuous or intermittent therapy for 96 weeks, showing that thrombocytopenia (<150 × 103 platelets/μl) was more prevalent in the intermittent therapy arm compared to the CT arm (25.4 vs. 9.8%, respectively, P < 0.001). Decreased platelet counts were correlated with changes in CD4 T-cell counts and plasma HIV-1 RNA levels (P < 0.001 for both) . This study had a relatively small sample size and used a very sensitive endpoint (thrombocytopenia defined as < 150 × 103 platelets/μl) that is clinically not thought of as thrombocytopenia. Whereas in the SMART study, the much larger sample size and the more accurate endpoint (thrombocytopenia defined as 100 × 103 platelets/μl) more convincingly showed the correlation between ART interruption and thrombocytopenia as well as allowing for the study of risk factors in more detail.
An elevated D-dimer reflects activation of the coagulation and fibrinolytic systems and is well known to be associated with increased risk of CVD and mortality from any cause [19–21]. Activation of the coagulation system may in certain conditions also cause thrombocytopenia, best described in syndromes such as disseminated intravascular coagulation, paroxysmal nocturnal haematuria , heparin-induced thrombocytopenia , thrombotic thrombocytopenic purpura and hemolytic uremic syndrome . These conditions are all characterized by endothelial and platelet activation as well as increased risk of thrombosis in spite of low platelet counts. Also in ITP, a paradoxically increased risk of thrombosis has been described . In-vivo platelet activation, circulating platelet-leukocyte-monocyte aggregates, platelet-derived microparticles, and immature reticulated platelets have all been implicated in the mechanism of a prothrombotic state in patients with ITP [26,27]. Reticulated platelets are immature platelets with increased mass and a greater prothrombotic potential compared with mature platelets [28–30]. In our study, we confirmed the inverse association between D-dimer, HIV-1 RNA levels, and platelet counts in the context of interrupting HIV treatment, suggesting that HIV-1 viremia might activate platelets and coagulation factors.
In the SMART study, reinitiation of ART was not associated with an improvement in inflammatory parameters (hsCRP, IL-6) . However, we have noted that platelet counts increased in most patients with HAART reintroduction, returning to their study entry levels. This lack of association does not support the hypothesis that unspecific inflammation is the sole force driving thrombocytopenia. As seen before, our data point to uncontrolled HIV-1 replication itself (most probable) or an activation in the coagulation pathways as pivotal underlying factors in the genesis of platelet count decreases.
Our study has some limitations that should be noted. Platelet levels were collected retrospectively at sites, only where available. Thus, we did not have platelet data for all SMART participants. By excluding sites that were unable to report platelet data on participants who had died or were lost to follow-up, we exclude 229 participants who experienced serious non-AIDS events. This resulted in our having limited power to study the effect of platelet kinetics on clinical event risk.
In summary, we have shown a significant decrease in platelet counts as well as a higher rate of thrombocytopenia (defined as <100.000/μl) after treatment interruption in SMART, which was related to increased viral replication and D-dimer levels. Most patients recovered to entry platelet levels after antiretroviral treatment reinitiation. Our data give support to uncontrolled HIV-1 replication and/or activation of coagulation pathways as the main factors underlying platelet count decreases.
We thank the participants who participated in SMART, the SMART study team (see below), and the INSIGHT Executive Committee.
The SMART Study Group: SMART was initiated by the Terry Beirn Community Programs for Clinical Research on AIDS (CPCRA) and implemented in collaboration with Investigators in international coordinating centers in Copenhagen (Copenhagen HIV Programme), London (Medical Research Council, Clinical Trials Unit), Sydney (National Centre in HIV Epidemiology and Clinical Research) and Washington (CPCRA). Participating staff are listed below.
Copenhagen International Coordinating Center: J.D. Lundgren, K.B. Jensen, D.C. Gey, L. Borup, M. Pearson, P.O. Jansson, B.G. Jensen, J. Tverland, H. Juncker-Benzon, Z. Fox, A.N. Phillips.
London Internatioal Coordinating Center: J.H. Darbyshire, A.G. Babiker, A.J. Palfreeman, S.L. Fleck, W. Dodds, E. King, B. Cordwell, F. van Hooff, Y. Collaco-Moraes.
Sydney International Coordinating Center: D.A. Cooper, S. Emery, F.M. Drummond, S.A. Connor, C.S. Satchell, S. Gunn, S. Oka, M.A. Delfino, K. Merlin, C. McGinley.
Washington International Coordinating Center: F. Gordin, E. Finley, D. Dietz, C. Chesson, M. Vjecha, B. Standridge.
INSIGHT Network Coordinating Center: J.D. Neaton, G. Bartsch, A. DuChene, M. George, B. Grund, M. Harrison, E. Krum, G. Larson, C. Miller, R. Nelson, J. Neuhaus, M.P. Roediger, T. Schultz.
ECG Reading Center: R. Prineas, C. Campbell, Z.-M. Zhang.
Endpoint Review Committee: G. Perez (co-chair), A. Lifson (co-chair), D. Duprez, J. Hoy, C. Lahart, D. Perlman, R. Price, R. Prineas, F. Rhame, J. Sampson, J. Worley.
NIAID Data and Safety Monitoring Board: M. Rein (chair), R. DerSimonian (executive secretary), B.A. Brody, E.S. Daar, N.N. Dubler, T.R. Fleming, D.J. Freeman, J.P. Kahn, K.M. Kim, G. Medoff, J.F. Modlin, R. Moellering Jr, B.E. Murray, B. Pick, M.L. Robb, D.O. Scharfstein, J. Sugarman, A. Tsiatis, C. Tuazon, L. Zoloth.
NIH, NIAID: K. Klingman, S. Lehrman.
SMART Clinical Site Investigators by country (SMART enrollment): Argentina (147): J. Lazovski, W.H. Belloso, M.H. Losso, J.A. Benetucci, S. Aquilia, V. Bittar, E.P. Bogdanowicz, P.E. Cahn, A.D. Casiró, I. Cassetti, J.M. Contarelli, J.A. Corral, A. Crinejo, L. Daciuk, D.O. David, G. Guaragna, M.T. Ishida, A. Krolewiecki, H.E. Laplume, M.B. Lasala, L. Lourtau, S.H. Lupo, A. Maranzana, F. Masciottra, M. Michaan, L. Ruggieri, E. Salazar, M. Sánchez, C. Somenzini.
Australia (170): J.F. Hoy, G.D. Rogers, A.M. Allworth, JStC Anderson, J. Armishaw, K. Barnes, A. Carr, A. Chiam, J.C.P. Chuah, M.C. Curry, R.L. Dever, W.A. Donohue, N.C. Doong, D.E. Dwyer, J. Dyer, B. Eu, V.W. Ferguson, M.A.H. French, R.J. Garsia, J. Gold, J.H. Hudson, S. Jeganathan, P. Konecny, J. Leung, C.L. McCormack, M. McMurchie, N. Medland, R.J. Moore, M.B. Moussa, D. Orth, M. Piper, T. Read, J.J. Roney, N. Roth, D.R. Shaw, J. Silvers, D.J. Smith, A.C. Street, R.J. Vale, N.A. Wendt, H. Wood, D.W. Youds, J. Zillman.
Austria (16): A. Rieger, V. Tozeau, A. Aichelburg, N. Vetter.
Belgium (95): N. Clumeck, S. Dewit, A. de Roo, K. Kabeya, P. Leonard, L. Lynen, M. Moutschen, E. O’Doherty.
Brazil (292): L.C. Pereira Jr, T.N.L. Souza, M. Schechter, R. Zajdenverg, M.M.T.B. Almeida, F. Araujo, F. Bahia, C. Brites, M.M. Caseiro, J. Casseb, A. Etzel, G.G. Falco, E.C.J. Filho, S.R. Flint, C.R. Gonzales, J.V.R. Madruga, L.N. Passos, T. Reuter, L.C. Sidi, A.L.C. Toscano.
Canada (102): D. Zarowny, E. Cherban, J. Cohen, B. Conway, C. Dufour, M. Ellis, A. Foster, D. Haase, H. Haldane, M. Houde, C. Kato, M. Klein, B. Lessard, A. Martel, C. Martel, N. McFarland, E. Paradis, A. Piche, R. Sandre, W. Schlech, S. Schmidt, F. Smaill, B. Thompson, S. Trottier, S. Vezina, S. Walmsley.
Chile (49): M.J. Wolff Reyes, R. Northland.
Denmark (19): L. Ostergaard, C. Pedersen, H. Nielsen, L. Hergens, I.R. Loftheim, K.B. Jensen.
Estonia (5): M. Raukas, K. Zilmer.
Finland (21): J. Justinen, M. Ristola.
France (272): P.M. Girard, R. Landman, S. Abel, S. Abgrall, K. Amat, L. Auperin, R. Barruet, A. Benalycherif, N. Benammar, M. Bensalem, M. Bentata, J.M. Besnier, M. Blanc, O. Bouchaud, A. Cabié, P. Chavannet, J.M. Chennebault, S. Dargere, X de la Tribonniere, T. Debord, N. Decaux, J. Delgado, M. Dupon, J. Durant, V. Frixon-Marin, C. Genet, L. Gérard, J. Gilquin, B. Hoen, V. Jeantils, H. Kouadio, P. Leclercq, J.-D. Lelièvre, Y. Levy, C.P. Michon, P. Nau, J. Pacanowski, C. Piketty, I. Poizot-Martin, I. Raymond, D. Salmon, J.L. Schmit, M.A. Serini, A. Simon, S. Tassi, F. Touam, R. Verdon, P. Weinbreck, L. Weiss, Y. Yazdanpanah, P. Yeni.
Germany (215): G. Fätkenheuer, S. Staszewski, F. Bergmann, S. Bitsch, J.R. Bogner, N. Brockmeyer, S. Esser, F.D. Goebel, M. Hartmann, H. Klinker, C. Lehmann, T. Lennemann, A. Plettenberg, A. Potthof, J. Rockstroh, B. Ross, A. Stoehr, J.C. Wasmuth, K. Wiedemeyer, R. Winzer.
Greece (95): A. Hatzakis, G. Touloumi, A. Antoniadou, G.L. Daikos, A. Dimitrakaki, P. Gargalianos-Kakolyris, M. Giannaris, A. Karafoulidou, A. Katsambas, O. Katsarou, A.N. Kontos, T. Kordossis, M.K. Lazanas, P. Panagopoulos, G. Panos, V. Paparizos, V. Papastamopoulos, G. Petrikkos, H. Sambatakou, A. Skoutelis, N. Tsogas, G. Xylomenos.
Funding: Support provided by National Institutes of Health grants (NIH Grants U01-AI068641, U01-AI046362, and U01-AI042170).
Authorship and disclosures: E.Z. and J.N. coordinated the research, supervised by J.D.L. E.Z., J.N., and J.D.L. wrote the first edition of the manuscript and J.V.B., C.S., J.M.L., A.P. and M.C. critically reviewed and revised it. The INSIGHT Scientific Steering Committee has also reviewed the manuscript.
Conflicts of interest
There are no conflicts of interest.
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Keywords:© 2013 Lippincott Williams & Wilkins, Inc.
antiretroviral therapy; D-dimer; HIV; platelets; strategies for management of antiretroviral therapy