**3. Thalidomide and thrombosis**

Thalidomide is a glutamic acid derivative that exerts potent anti-angiogenic and immunomodulatory activity and has revolutionized clinical management of patients with myeloma. Thalidomide is effective in relapsed or refractory and newly diagnosed MM. However, from experience with myeloma patients, VTE has recently emerged as the single most important complication. The anti-myeloma effect is mediated by several mechanisms in myeloma cells directly as well as by the microenvironment [11]. Thalidomide induces G1 growth arrest/apoptosis by inhibiting NF-ΚB [12] and activating caspase. It inhibits adhesion of myeloma cells to bone marrow stromal cells, and inhibits secretion of cytokines (VEGF, [13], βFGF, [14-16], HGF, [17], TNFa, [18], IL-6, [19] and soluble IL-6 receptor (sIL-6R) [20]); up-regulate ICAM-1, [21] VCAM-1, IL-10, [22-23] and IL-12, [24]. Finally, it induces T cell and NK cell anti-myeloma immunity and inhibits angiogenesis [25].

The occurrence of VTE with the use of thalidomide was reported for the first time by Osman and Rajkumar in two independent phase 2 trials [31] in 2001. The combination of thalidomide (100–200 mg/day), doxorubicin (36 mg/m2 on the first day of each 28-day cycle,) and dexamethasone (40 mg daily on days 1–4, 9–12, and 17–20 of each cycle) produced symptomatic deep venous thrombosis in four of the first 15 enrolled patients (27%) and the trial was therefore stopped. The other phase 2 trial combined thalidomide and dexamethasone for the treatment of patients with newly diagnosed myeloma at the Mayo Clinic. Seven percent (3/45) had thrombotic events. Cavo [32] and Rajkumar [33] in another Pathophysiology and Clinical Aspects of 116 Venous Thromboembolism in Neonates, Renal Disease and Cancer Patients

MGUS, and MM, respectively [7]. The incidence of VTE in MM patients is difficult to estimate and varies from 3–10%. On the other hand, the recent introduction of the antimyeloma therapy class called immunomodulatory drugs (IMiDs), substantially increased

The exact pathogenesis of thrombosis in plasma cell dyscrasia is multifactorial and poorly understood. Prothrombotic coagulation abnormalities are found in patients with newly diagnosed multiple myeloma, including elevated levels of von Willebrand antigen factor, factor VIII and tissue factor, as well as decreased protein S and thrombosposndin [8]. Proinflammatory and angiogenic cytokines such as interleukin-6, tumor necrosis factor and vascular endothelial growth factor (VEGF) are elevated in MM and could activate the coagulation system [9]. A recently described mechanism of hypercoagulability in cancer patients, including MM patients, is acquired activated protein C resistance (APC-R). APC-R, in the absence of factor V Leiden mutation, was present in almost one-quarter of newly diagnosed myeloma patients and significantly increased the risk of VTE [10]. The possible production of auto-antibodies against protein C in these patients could explain the transient APC resistance phenotype. However, to date, we know of no single prothrombotic abnormality that can be used to predict which patients with plasma cell dyscrasia will develop VTE. Other risk factors of MM-associated thrombosis are involved, such as older age, immobility, prior or family history of VTE and the presence of other medical comorbidities, immobility due to pain and/or surgery, indwelling central venous catheters, extrinsic venous compression by plasmacytomas, and the presence of inherited factors such as factor V Leiden. However, the dominant risk factor for VTE in MM is the type of drug

Thalidomide is a glutamic acid derivative that exerts potent anti-angiogenic and immunomodulatory activity and has revolutionized clinical management of patients with myeloma. Thalidomide is effective in relapsed or refractory and newly diagnosed MM. However, from experience with myeloma patients, VTE has recently emerged as the single most important complication. The anti-myeloma effect is mediated by several mechanisms in myeloma cells directly as well as by the microenvironment [11]. Thalidomide induces G1 growth arrest/apoptosis by inhibiting NF-ΚB [12] and activating caspase. It inhibits adhesion of myeloma cells to bone marrow stromal cells, and inhibits secretion of cytokines (VEGF, [13], βFGF, [14-16], HGF, [17], TNFa, [18], IL-6, [19] and soluble IL-6 receptor (sIL-6R) [20]); up-regulate ICAM-1, [21] VCAM-1, IL-10, [22-23] and IL-12, [24]. Finally, it

induces T cell and NK cell anti-myeloma immunity and inhibits angiogenesis [25].

The occurrence of VTE with the use of thalidomide was reported for the first time by Osman and Rajkumar in two independent phase 2 trials [31] in 2001. The combination of thalidomide (100–200 mg/day), doxorubicin (36 mg/m2 on the first day of each 28-day cycle,) and dexamethasone (40 mg daily on days 1–4, 9–12, and 17–20 of each cycle) produced symptomatic deep venous thrombosis in four of the first 15 enrolled patients (27%) and the trial was therefore stopped. The other phase 2 trial combined thalidomide and dexamethasone for the treatment of patients with newly diagnosed myeloma at the Mayo Clinic. Seven percent (3/45) had thrombotic events. Cavo [32] and Rajkumar [33] in another

risk of VTE in multiple myeloma.

administered.

**3. Thalidomide and thrombosis** 

two phase 3 trials confirmed the preliminary observations of increased incidence of VTE in newly diagnosed MM treated with thalidomide plus dexamethasone (16 and 17% of VTEs). (*table 1 below*).


RR: refractory / relapsed; ND: newly diagnosed.

Table 1. Incidence of thalidomide-associated venous thromboembolism without VTE prophylaxis

Similarly, when thalidomide was combined with melphalan and steroids, the incidence of VTE was 9–20% in newly diagnosed elderly patients [36-38]. In all of these studies, the major risk of thrombosis occurs early after initiation of the treatment, when the tumor load is maximal. Thus, this complication may be related to the release of thrombogenic factors from myeloma cells rather than to cumulative drug exposure [49].

Thrombosis Associated with Immunomodulatory Agents in Multiple Myeloma 119

The majority of thrombotic events described in patients receiving treatment with thalidomide have been venous, but occasional arterial thrombotic events have also been reported [60-63]. In a prospective analysis of arterial thrombosis risk, the incidence of arterial thrombosis in patients with newly diagnosed MM treated with three cycles of thalidomide, doxorubicin and dexamethasone (TAD group) was 4.5%. However, the true incidence of arterial thrombosis could have been underestimated in the TAD group due to prophylactic use of LMWH. High factor VIII:C levels, possibly reflecting disease activity, could contribute to the risk of arterial thrombosis especially in patients with known

Lenalidomide, a more potent immunomodulatory derivative of thalidomide, was designed to increase the anti-myeloma efficacy of thalidomide, while possibly reducing side effects like neuropathy and thrombosis. However, although neuropathy is not an important lenalidomide-related side effect, thrombosis continues to be one of the most important side effects, especially when lenalidomide is given in combination with dexamethasone or

Regimen n Status of disease Incidence (%)

Richardson, 2009 65 222 RR 4

 Dimopoulos, 2007 66 176 RR 11.4 Weber, 2007 67 177 RR 14.7 Zonder, 2005 68 38 ND 75 Rajkumar 2010 69 223 ND 26

Rajkumar, 2010 69 222 ND 12

Richardson, 2010 70 66 ND 6

Table 2. Incidence of lenalidomide-associated venous thromboembolism without VTE

In two large phase 3 trials comparing lenalidomide plus dexamethasone to dexamethasone alone without mandated thromboprophylaxis in patients with RRMM, the incidences of VTE in the LD arm were 11.4 and 14.7%, compared with 4.6 and 3.4% in the DEX alone arm [66-67]. The incidence was even higher in patients with newly diagnosed MM (up to 75%) who were treated with lenalidomide plus dexamethasone [68]. In all these trials conducted in RR and newly diagnosed MM patients, dexamethasone was given at high dose (three pulses of 40 mg for 4 days, total amount per cycle: 480 mg). Interestingly, the rate of VTE was significantly lower when lenalidomide was combined

cardiovascular risk factors [64].

chemotherapy (*table 2*).

**Lenalidomide in monotherapy** 

**Lenalidomide / Dexamethasone** 

**Lenalidomide / Dexamethasone /** 

**Bortezomib** 

prophylaxis

**Lenalidomide / Dexamethasone Low Dose** 

RR: refractory / relapsed; ND: newly diagnosed.

**4. Lenalidomide and thrombosis** 

A meta-analysis of studies of thalidomide in MM, which involved 3,322 patients, showed that patients receiving thalidomide were 2.1 times as likely to have a VTE event compared with those who were not receiving thalidomide (p<0.01). Those receiving thalidomide plus dexamethasone were 3.1 times as likely to have a VTE event (p<0.01), and those receiving thalidomide in addition to other chemotherapy agents were 1.5 times as likely to have a VTE event (p<0.01) [2].

The pathogenesis of thalidomide-associated thrombosis has not yet been established. Zangari et al. [10] tested for hypercoagulability in 62 newly diagnosed MMs, and found that DVT was more frequent in those patients with acquired APC resistance (36 *vs.* 15%, p<0.04). The pre-existing elevated factor VIII coagulant activity and von Willebrand factor antigen have also been related with thalidomide-associated thrombosis [50]. In an experimental model, Kausahal et al. demonstrated that the addition of thalidomide to uninjured arterial endothelium did not cause any appreciable change, whereas thalidomide added to adriamycin-injured (8–24 h) endothelial cells resulted in endothelial dysfunction by altering the expression of PAR-1 in injured endothelium [51].

Cases of VTE that began shortly after the initiation of treatment with recombinant human erythropoietin in patients who had been receiving thalidomide for some time have been reported [52]. However, another study found no apparent increased risk of thrombosis in 199 cases of myeloma given thalidomide with or without erythropoietin. Of the 49 patients receiving both drugs, 8.1% developed thrombosis compared with 9.3% of the 150 patients on thalidomide who did not receive erythropoietin [53].

The genetic susceptibility to developing a VTE in response to thalidomide therapy has been also evaluated. The lack of a strong association with genetic variations in the coagulation cascade, such as factor V Leiden or G20210A prothrombin mutation, suggests that VTE risk is mediated via alternative mechanisms. Johnson et al [54] identified 18 SNPs, using a custom-built molecular inversion probe (MIP)-based single nucleotide polymorphism (SNP) chip. There were two "response to stress" groups: a response to DNA damage group, including *CHEK1*, *XRCC5*, *LIG1*, *ERCC6*, *DCLRE1B* and *PARP1*, and a cytokine response group containing *NFKB1*, *TNFRSF17*, *IL12B* and *LEP*. A third apoptosis-related group with *CASP3*, *PPARD* and *NFKB1* was also found.

Interestingly, no thromboembolic events were observed in a group of 30 patients with relapsed MM treated with a bortezomib, melphalan, prednisone and thalidomide (VMPT) combination despite the absence of any anticoagulant prophylaxis [55]. A recent review of phase 3 trials of bortezomib- and/or IMiD-based therapy in frontline MM, together with other studies of novel combination regimens concluded that bortezomib-based regimens were typically associated with DVT/PE rates of ≤ 5%, similar to those seen with melphalanprednisone and dexamethasone, whereas IMiD-based bortezomib-free regimens were generally associated with higher rates [56]. These results suggested the existence of a protective effect of coadministration of thalidomide or lenalidomide with bortezomib [57- 58]. Zangari et al prospectively described *in vivo* effects of bortezomib from routine tests of blood coagulation and platelet function in treated MM patients. This pilot clinical trial showed *in vivo* that even a short exposure to bortezomib can affect platelet function. Platelet aggregation was reduced after bortezomib infusion with most of the commonly agonists used (ADP, epinephrine and ristocetin) on both days of treatment [59].

The majority of thrombotic events described in patients receiving treatment with thalidomide have been venous, but occasional arterial thrombotic events have also been reported [60-63]. In a prospective analysis of arterial thrombosis risk, the incidence of arterial thrombosis in patients with newly diagnosed MM treated with three cycles of thalidomide, doxorubicin and dexamethasone (TAD group) was 4.5%. However, the true incidence of arterial thrombosis could have been underestimated in the TAD group due to prophylactic use of LMWH. High factor VIII:C levels, possibly reflecting disease activity, could contribute to the risk of arterial thrombosis especially in patients with known cardiovascular risk factors [64].
