Drug-induced thrombocytopenia – etiology and alternative therapeutic approaches

Introduction and aim . The cumulative incidence of drug-induced thrombocytopenia (DIT) is 10 cases per one million people per year with a prevalence of approximately 25% in critically ill patients. This review provides a comprehensive view of drug-induced thrombocytopenia, diagnosis, underlying mechanisms, common strategies in therapeutics, and potential alternatives. Material and methods . Databases such as “Google Scholar”, “PubMed”, “Medline” and “MDPI” was used for literature review with the keywords, “platelets”, “platelet disorders”, “thrombocytopenia”, “drug-induced”, “oxidative stress” “plant extracts”, “phy-tochemicals”, “antioxidants”, for the articles published between 2013-2023 and written in the English language. Analysis of the literature . Several antimicrobials, anti-cancer drugs, and antivirals are often reported to cause adverse effects during treatment, such as thrombocytopenia. A thorough understanding of the underlying pathophysiology is important for appropriate treatment. Even though an improvement in platelet count is observed after the discontinuation of the causative drug, there is a dire need for treatment in some cases due to associated complications. There are various pitfalls with conventional treatments which include clinical complications and lack of effectiveness. Conclusion . Interventions in therapeutics through antioxidants can aid in faster recovery. Various plant extracts and phyto-chemicals have been employed as therapeutics in platelet disorders due to their exceptional antioxidant activity. It is imperative to explore the bioactive components of natural products and their influence on platelet efficacy. Also, it highlights how antioxidants can be used as a safe, yet effective option as therapeutics for treating a complicated disorder such as DIT or be used as supplements to prevent adverse effects of existing treatments involving antibiotics and chemotherapeutics.


Introduction
Platelets or thrombocytes are blood cells that are continuously produced from megakaryocytes in a process called Thrombopoiesis, mainly in the bone marrow.These anucleate cells range from ~150,000-450,000/µL in healthy individuals. 1They are found circulating in the blood for 8-10 days and are eliminated in the spleen and liver. 2 The main function of the platelet is to react to vascular damage and form a blood clot at the site to stop bleeding. 3Nonetheless, platelets are also involved in several other functions such as affecting tumor pro-gression by stimulating angiogenesis, separating the lymphatic circulation during embryogenesis, and helping to close ductus arteriosus at the time of birth.They are a significant part of primary immunity and contribute inflammatory mediators. 4latelet disorders are primarily due to following reasons: (i) an increase in platelet number, (ii) a decrease in platelet numbers, or (iii) platelet dysfunction. 5These can lead to either defective hemostatic plug formation leading to bleeding or spontaneous clot formation leading to thrombosis. 6latelet disorders are classified into: i. Platelet function disorders: Inherited or acquired, due to loss or gain of platelet function, change in size and morphology of the platelets, and associated with clinically important bleeding defects.7 ii.High platelet count conditions or Thrombocytosis: Abnormal accumulation of platelets in the blood is further classified into primary or essential and secondary or reactive thrombocytosis. 8,9ii.Low platelet count conditions or Thrombocytopenia: Atypically low platelet levels (under 20,000 per mm 3 ) due to an underlying cause. 10

Aim
This review provides a comprehensive view of drug-induced thrombocytopenia, its diagnosis, underlying mechanisms, common strategies in therapeutics, and potential alternatives such as phytochemicals.

Thrombocytopenia
Thrombocytopenia is characterized by a massive reduction in platelets in the peripheral blood, caused by a myriad of both congenital and acquired causes.Acute and severe decrease in platelet count exposes the individuals to an increased risk of developing spontaneous hemorrhage or thrombosis.Hemostasis entirely depends upon the functioning and the number of platelets in circulating blood and any deviation in the number can cause many complications. 11he major causes of thrombocytopenia can be summarized as follows: 12,13 a. Decrease in differentiation of hematopoietic stem cells in bone marrow b.Impairment of megakaryocyte maturation c.Disruption of endothelial adhesion of megakaryocytes d.Increase in peripheral platelet destruction due to immune and non-immune mediated pathways e. Increase in splenic sequestration f.Accelerated clearance from the circulation g.Oxidative Stress

Drug-induced thrombocytopenia (DIT)
The main threat in any treatment procedure is the side effects caused by therapeutic drugs.Drugs administered for the treatment of a clinical symptom are capable of causing a different disorder altogether, based on their mode of action at the molecular level.DIT is one such disorder with a cumulative incidence of about 10 cases per million population per year and a prevalence as high as 25% in critically ill patients. 14DIT is often underdiagnosed and underreported and puts the patients at an increased risk of hemorrhage.
Studies have been conducted during the past two decades based on the types of drugs that specifically induce thrombocytopenia.Around 300 drugs have been clinically implicated in causing DIT as an adverse effect during the treatment procedure.Commonly reported drugs can be categorized into antibiotics, chemotherapeutic drugs, steroids and cardiovascular drugs.Chemotherapy regimens involving dexamethasone + cytarabine + cisplatin, isophosphamide + carboplatin + etoposide, gemcitabine + dexamethasone + cisplatin and gemcitabine + and oxaliplatin are most commonly reported to cause DIT in cancer patients. 15Some of the glycoprotein IIb/IIIa (GPIIb/IIa) inhibitors have also been reported in DIT cases. 14,16Drug-related disorders are common in small molecules as well as biotherapeutics.Although they were believed to be safe due to their specific action, many biotherapeutic drugs have been reported to lead to adverse reactions leading to loss in blood cell count and functions. 16Some of the clinically important cases of DIT are listed in Table 1.
A detailed history of the patient and timely diagnosis is necessary to understand the underlying etiology.However, DIT often manifests in two ways: a.One to two weeks after beginning a new drug or suddenly, after a single dose when the drug was previously taken intermittently. 22b.Immediately after the first administration of antithrombotic agents that block fibrinogen binding to platelet GP IIb-IIIa. 14,37IT varies extensively from case to case and is often misdiagnosed as immune thrombocytopenic purpura and patients receive inappropriate treatment, leading to several complications.There have been cases where patients may not consider that self-regulated medications, beverages, foods, or herbal remedies are relevant to their bleeding symptoms and may not report them. 38horough understanding of the underlying pathophysiology plays a crucial role in devising a treatment of DIT.Identification of the trigger for the sudden fall in platelet count is the first and foremost step in the diagnosis of DIT.Secondly, excluding pseudothrombocytopenia is critical which occurs due to in vitro factors such as anticoagulants used during blood collection.][41] Mechanisms Majorly, the mechanisms can be classified into (i) platelet destruction and (ii) platelet production.i. Platelet destruction: Platelet clearance due to increase in platelet desialylation by NanA neuraminidase and the Ashwell-Morell receptor (AMR)-dependent pathway can be the major causes of DIT during infection. 42Cross-Reaction of the drugs with endogenous thrombopoietin can lead to the neutralization of platelets in the blood stream. 43Off-target platelet binding and activation due to the overloading of the therapeutic drug in blood can result in platelet acti-vation and eventually destruction. 44The immune system can also play a major role in platelet destruction by inducing antiplatelet antibodies against the murine sequences of the chimeric Fab molecule. 31i.Platelet production: Low platelet levels may be due to the inhibition of thrombopoietin signaling in early megakaryokinesis leading to decreased platelet production. 45Myelosuppression/toxicity by targeting the epidermal growth factor receptor pathway, vascular endothelial growth factor (VEGF) pathway, and BCR-ABL pathway can contribute to low platelet count as there is an abnormality at the production level or induction of platelet apoptosis. 37,46,47Bone marrow toxicity caused by some drugs can inhibit megakaryocyte differentiation and maturation which can impact platelet numbers. 48Platelet consumption can also occur due to endothelial dysfunction from lack of VEGF or suppression of hematopoiesis. 49Table 2 outlines various mechanisms that reduce platelet numbers based on different classes of drugs and their target pathways.

Bone marrow toxicity and inhibits megakaryocyte differentiation and maturation
Bevacizumab 49 Platelet consumption due to endothelial dysfunction from lack of VEGF Anti-CD40L mAbs 50 Inhibits disaggregation of ADP-aggregated platelets Abciximab 51 Induction of antiplatelet antibodies directed against the murine sequences of chimeric Fab molecule TNF-α inhibitors 52,53 Induces autoantibodies including antiplatelet antibodies or suppression of hematopoiesis AMG X and LY2541546 54,55 Platelet activation by direct binding to platelets and/ or megakaryocytes causes the release of serotonin Thrombopoietic agents-rHu-TPO or PEG-rHuMGDF 43 Cross-Reacts with endogenous thrombopoietin and neutralizes CH12 humAb 44 Off-target platelet binding and activation resulting in platelet destruction

Treatments
Treatment strategies for DIT focus primarily on the recovery of lost platelets.In most cases, physicians identify the drug responsible for the condition and take the first step by switching the suspected drug to an alternate drug.Cessation of administering a suspected drug generally reverses the condition of DIT in most cases. 14However, treatment is required in cases where the therapeutic drug is crucial for the patient or when patients experience life-threatening bleeding.Conventional treatment is advised based on the following strategies outlined below.

Common approaches
The outline of commonly used strategies in the treatment of DIT is summarized in Fig. 2 and can be explained as follows: Nonetheless, utilizing platelets stored in blood banks having limited shelf-life of up to 5 days can be ineffective due to the effect of storage lesions. 61,62v. Surgical: Splenectomy is a surgical procedure that partially or completely removes the spleen to increase the platelet count.This procedure aids in the release of splenic sequestered platelets into circulation, thereby instantly elevating the platelets.However, the major drawback of the procedure is that it may adversely affect portal hypertension.Therefore, portal decompression is more preferred to improve thrombocytopenia.Nonetheless, surgical procedures are the least recommended for DIT as it is mostly unnecessary and may cause further damage during the procedure. 63vi.Other drugs: Thrombopoietin receptor (TPO-R) agonists are potential therapeutic drugs for DIT which stimulates the platelet production in the bone marrow.These act as replacement for thrombopoietin and elevate the platelet count.Spleen tyrosine kinase (SYK)-inhibitors are yet another approved class of drugs for DIT which can reduce antibody-mediated platelet destruction by immunosuppression mechanism. 64,65Some of the examples for drugs include romiplostim, eltrombopag, imiglucerase, avatrombopag and fostamatinib. 58

Alternative approaches
Oxidative stress in cells and tissues due to reactive oxygen species (ROS) is attributed as a primary causative agent in several disease conditions.Free radicals are highly unstable molecules and are highly reactive, thereby attacking the molecules in a biological system, causing metabolic malfunction, and cellular protein destruction, and can cause cell death. 66,67ntioxidants scavenge ROS and repair oxidative damage.Primary antioxidants scavenge ROS by hydrogen atom transfer (HAT) mechanism, by donating an H-atom, or act by single electron transfer (SET) mechanism.The secondary antioxidants neutralize the ROS using prooxidant catalysts by quenching free radicals and are thus exhausted. 68Plant extracts and phytochemicals have excellent antioxidant capacity and are explored to reduce the possibility of the occurrence of disorders.There are reports that antioxidants alleviate the condition when used as an alternative therapeutic strategy or taken as a supplement or in numerous disease conditions. 69,70tudies have proven that various plant extracts can be used as therapeutic agents in the treatment of thrombocytopenic conditions.These plant extracts have been routinely tested in animal models for their effectiveness in stimulating the production of platelets in thrombo-cytopenic condition.This ability of plant extracts is attributed to their antioxidant capacity.There are many studies dedicated to the identification of the bioactive compound in such extracts.Most bioactive components are phenolic acids, polyphenols, flavonoids or alkaloids in nature.The major challenge lies in identifying the specific bioactive compound in these extracts.Also, their influence on platelet functions is extremely under-reported.Nevertheless, specific antioxidants have been proven to be effective in improving platelet functions such as aggregation, granular secretion, activation and apoptosis and increasing the ability of endogenous antioxidant defense. 71,72Our preliminary studies have indicated that antioxidants such as L-carnitine and vanillic acid are beneficial for platelet survival. 73,74Table 3 and Table 4 provide details of plant extracts and antioxidants, respectively used under different thrombocytopenic conditions to improve platelet count.These studies are critical to categorize the antioxidants into either platelet agonists or anti-platelet agents as they are applied under rather entirely contrasting pathophysiological circumstances.However, there are a few studies on using antioxidants and herbal products as supplements in human subjects with clinically significant thrombocytopenic conditions.3][94] Effect of some antioxidants on platelet functions can be dose-dependent.They act as platelet agonists at higher concentration and can become anti-platelet at lower concentrations. 95Hence, it is essential to evaluate such antioxidants and optimize the dose for use as supplements in alternate therapeutics.

Conclusion
Alternate therapies using phytochemicals can aid in faster recovery and reduce mortality due to their remarkable antioxidant properties.Plant extracts have shown promising results in increasing the number of platelets in patients with thrombocytopenia.Although these natural remedies are not a substitute for conventional therapies, they can be used a supplementary therapy to improve the effectiveness of treatments.Based on the studies conducted to this point, they can also be used as supplements to prevent adverse effects of existing treatments involving antibiotics and chemotherapeutics.Antioxidant therapies are currently followed for various clinical conditions and are a promising arena for further exploration.However, these therapies are still in its infancy, and have gained attention in the recent past due to the awareness and benefits of traditional approaches.Several scientific reports encourage using antioxidants as alternate therapeutic substances.Furthermore, antioxidants are investigated only in terms of enhancement of platelet count for DIT, but it is imperative to study their effect on platelet functions.Nonetheless, these antioxidants should be extensively studied for their mechanistic properties and physiological interactions to be regarded as one of the standard therapeutic strategies.

Fig. 2 .
Fig. 2. Therapeutic strategies for Drug-induced thrombocytopenia Aspirin-induced thrombocytopeniaBoth plant extracts possess plateletincreasing property in combination or alone

Table 1 .
Cases of drug-induced thrombocytopenia and treatments

Table 2 .
Mechanisms of the drugs inducing thrombocytopenia

Table 3 .
Plant extracts as potential therapeutics

Table 4 .
Antioxidants as potential therapeutics for thrombocytopenia