Published OnlineFirst June 28, 2011; DOI: 10.1158/1535-7163.MCT-11-0042
Molecular Cancer Review Therapeutics
Novel Therapeutic Options in Anaplastic Large Cell Lymphoma: Molecular Targets and Immunological Tools
Olaf Merkel1, Frank Hamacher1, Eveline Sifft1, Lukas Kenner2,3, and Richard Greil1 for the European Research Initiative on Anaplastic Large Cell Lymphoma
Abstract Anaplastic large cell lymphoma (ALCL) is a CD30-positive, aggressive T-cell lymphoma, and about half of the patients with this disease harbor the t(2;5)(p21;q35) translocation. This chromosomal aberration leads to fusion of the NPM gene with the ALK tyrosine kinase, leading to its constitutive activation. To date, treatment options include polychemotherapy (e.g., cyclophosphamide, doxorubicin, vincristine, and prednisone), which is sometimes combined with radiation in the case of bulky disease, leading to remission rates of 80%. However, the remaining patients do not respond to therapy, and some patients experience chemo-resistant relapses, making the identification of new and better treatments imperative. The recent discovery of deregulated ALK in common cancers such as non–small cell lung cancer and neuroblastoma has reinvigorated industry interest in the development of ALK inhibitors. Moreover, it has been shown that the ALK protein is an ideal antigen for vaccination strategies due to its low expression in normal tissue. The characterization of microRNAs that are deregulated in ALCL will yield new insights into the biology of ALCL and open new avenues for therapeutic approaches in the future. Also, CD30 antibodies that have been tested in ALCL for quite a while will probably find a place in forthcoming treatment strategies. Mol Cancer Ther; 10(7); 1127–36. 2011 AACR.
Introduction showing large cells with abundant cytoplasm and eccentric, lobulated nuclei (3). However, the clear dis- Anaplastic large cell lymphoma (ALCL) is a rare, tinction of ALCL from other disease entities was made aggressive, mature T-cell lymphoma that affects both possible by the work of Stein and colleagues (4), who children and adults. ALCL accounts for 10% to 15% of showed that the surface antigen CD30 is expressed by all non-Hodgkin’s lymphoma (NHL) cases in children ALCL cells. In about half of the patients, the translocation and 2% to 8% of NHL cases in adults. Clinically, ALCL t(2;5)(p23;q35) can be found, resulting in the expression of can be distinguished as a systemic disease or a localized the nucleophosmin-anaplastic lymphoma kinase (NPM- þ primary cutaneous entity. Systemic ALCL is a very ALK) fusion protein. ALK lymphomas represent 50% to aggressive lymphoma that involves different extranodal 85% of systemic ALCL cases and occur mainly in the first secondary sites, including soft tissue, skin, bone, lungs, 3 decades of life, predominantly in males. ALK ALCL and liver (1). Primary cutaneous ALCL accounts for <2% occurs in older patients, with a peak at 60 years of age and of the disease frequency but has a 5-year disease-free lower male predominance. ALK ALCL patients have a þ survival rate of 90% upon standard treatment (2). The less favorable prognosis than ALK ALCL patients after primary systemic and cutaneous subtypes have a similar treatment with polychemotherapy. Other fusion partners histologic appearance, with so-called hallmark cells of the ALK kinase, such as tropomyosin-3, 5-aminoimi- dazole-4-carboxamide ribonucleotide formyltransferase/ IMP cyclohydrolase, clathrin heavy chain, and TRK fused Authors' Affiliations: 1Laboratory for Immunological and Molecular Can- gene 1, have been described (Fig. 1). In most cases, the cer Research, Third Medical Department of Hematology, Medical Oncol- fusion partners contain a dimerization domain that leads ogy, Hemostaseology, Rheumatology and Infectiology, Paracelsus to constitutive activation of the ALK kinase (5). NPM is a Medical University Salzburg, Salzburg, Austria; 2Department of Clinical Pathology, Medical University Vienna, and 3Ludwig Boltzmann Institute for multifunctional 23 kDa protein that is involved in diverse Cancer Research, Vienna, Austria processes such as ribosome biogenesis, regulation of cell Note: The website for the European Research Initiative on Anaplastic division, and DNA repair. ALK is a trans-membrane Large Cell Lymphoma is www.erialcl.net receptor tyrosine kinase (RTK) of the insulin receptor Corresponding Author: Olaf Merkel, Laboratory for Immunological and superfamily (6). The normal function of ALK is still Molecular Cancer Research, Third Medical Department, Paracelsus Med- ical University Salzburg, Mullner€ Hauptstrasse 48, 5020 Salzburg, Austria. elusive, especially because knockout mice have only very Phone: 43-662-4482-1551; Fax: 43-662-4482-1570; E-mail: subtle phenotypes. In normal tissue, ALK expression has [email protected] mainly been found in the neonatal brain, peripheral doi: 10.1158/1535-7163.MCT-11-0042 nervous system, and spinal fluid, suggesting a function 2011 American Association for Cancer Research. in neuronal development.
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Toxin
SGN-35
CD30 TFG1 Figure 1. Sites of action of current and future ALCL therapeutics. The CLTC TM ALK kinase can be activated either NSCLC EML4 1 1620 aa by fusion to other proteins that lead to dimerization and thereby TK- ALCL NPM + ALK domain constitutive activation (ALCL and IMT), or by point mutations within IMT RanBP2 the full-length protein (NB). Both TPM3 Break point are amenable to ALK-TK inhibition. Another approach is to ATIC inhibit one of the many ALK- ALK activation activated downstream pathways, such as PI3K/Akt1/mTOR (7), Fusion Mutation JAK/Stat3 (8), and RAS/ERK (78). Several inhibitors are known to act ALCL NB directly on members of these pathways [e.g., PF-956980, JAK3 hsp 1058 1620 1 391 480 1030 1122 1620 aa inhibitor; S3I-201, STAT3 inhibitor; hsp 70 90 ABT-737 and ABT-263, BH-3 TK- mimetics inhibiting Bcl-2 and NPM ALK NH2 ALK COOH domain Bcl-xl; GGTI-298 (geranylgeranyltransferase 17-AAG 26 401 635 1058 1376 Geldanamycin inhibitor-298) and FTI (farnesyltransferase inhibitor), Ras ALK-tyrosine kinase inhibitors inhibitors; rapamycin and temsirolimus, mTOR inhibitors; and several Akt inhibitors]. In contrast, for ALCL patients PF-956980 without the ALK translocation, other treatment options would JAK3 GGTI-298 Pl3K S31-201 have to be considered. One such JNK FTI option may be the MMAE-coupled RAS Bcl-xl STAT3 Akt1/2 CD30 antibody SGN-35 (Seattle Mcl-1 JUN Akt-Inhibitors Genetics). Recently, we Bcl-2 Cyclins AP1 mTOR ERK1/2 Rapamycin characterized oncogenic miRNAs Temsirolimus that are specifically ABT-737 þ miR-29b Myc overexpressed in ALCL, ALK ABT-263 miR-101 (miR-17–92 cluster), or ALCL, ALK (miR-155), as well as miRNAs that have low expression in both forms of ALCL (e.g., miR- ALCL miR-17 29b and miR-101). These may 20a represent novel molecular targets miR-155 miR-155 20b for future ALCL therapies. ALK+ ALK- 93 106a
The fusion of the N-terminal part of NPM to the kinase patients often lacking the NPM-ALK translocation, may domain of ALK results in its constitutive activation. This influence the perception of a more negative outcome for leads to the activation of multiple downstream pathways, ALK negative patients. In patients older than 40 years, a þ including PI3K/Akt/mTOR (7), JAK/Stat3 (8), c-myc (9), prognostic difference between the ALK and ALK var- and cJun/JunB (ref. 10; Fig. 1). Mouse models that express iants of ALCL has not been found (13). In the most recent the NPM-ALK within the hematopoietic system have World Health Organization classification of this disease, þ shown the transforming capacity of this fusion protein the ALK and ALK subgroups were classified as 2 (10–12). However, it remains a mystery why the ALCL different disease entities, mainly due to their different variant that does not bear the translocation has the worst prognostic properties (14). The molecular features of prognosis. The skewed age distribution, with adult ALCL ALK-negative ALCL are largely unknown. Interest in
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Novel Drugs for ALCL
ALK kinase activation and its functional consequences from Streptomyces staurosporeus. UCN-01 was reported to increased tremendously after the recent finding that the have antitumor activity in a single patient with ALK- echinoderm microtubule-associated protein-like 4-ALK positive ALCL that was refractory to conventional CHOP kinase fusion protein (EML4-ALK) can be detected in therapy (27). Derivatives of staurosporine include the >6% of non–small cell lung cancer (NSCLC) patients (15). inhibitors CEP-14083 and CEP-14513 (Cephalon, Frazer, Soda and colleagues (16) developed a mouse model for PA), whose clinical use is hampered by unfavorable EML4-ALK–driven lung cancer and showed that ALK physical properties such as low solubility and stability inhibitors can be effectively used for treatment in the (28, 29). Second-generation inhibitors, such as compound murine system. By the end of 2009, Pfizer had launched a 18 (Cephalon, Frazer, PA), showed improved properties phase 3 clinical trial evaluating ALK inhibitors for the in vivo with oral bioavailability and effective inhibition of treatment of NSCLC bearing the EML4-ALK fusion. In tumor growth in the SCID mouse Sup-M2 tumor xeno- patients with neuroblastoma (NB), point mutations were graft model (30). Another class of naturally occurring found that resulted in constitutive ALK activation in 6% ALK modulators consists of 17-allylamino-17-demethox- to 12% of the patients (17–20). Recently, the EML4-ALK ygeldanamycin (17-AAG), geldanamycin, and herbimy- translocation was found with the use of exon array cin A. These modulators lead to indirect ALK inhibition profiling in subsets of breast and colorectal cancer (21). via binding to heat shock protein 90 (HSP-90), thereby þ ALK patients are invariably treated with polyche- enhancing the proteasome-mediated degradation of the motherapy—in most cases with cyclophosphamide, dox- ALK protein (31, 32). Other inhibitor classes include orubicin, vincristine, and prednisone (CHOP), which pyridine, thiazole, aminopyridine, diaminopyridine, causes complete remission in 80% of patients. Approxi- and fused ring systems. Li and colleagues (33) identified mately 60% of ALK patients only enter remission fol- a series of novel pyridones as kinase inhibitors of ALK by lowing CHOP treatment, leaving room for improvement stepwise in vitro screening followed by iterative template of treatment strategies. modification and computational ranking. One substance in this inhibitor family is the Novartis compound NVP- ALK Kinase Inhibitors TAE684, which inhibits proliferation of Karpas-299 and SU-DHL-1 cell lines with an IC50 range of 2 to 5 nM, The ALK cDNA encodes a protein of 177 kDa. The hindering autophosphorylation of NPM-ALK (34). This protein contains an extracellular ligand-binding domain, compound is highly NPM-ALK specific, as the activity of a transmembrane-spanning region, and a cytoplasmic other insulin receptor kinases is not significantly kinase catalytic part (the TK domain), which has a major impaired. Another member of this inhibitor class is the autophosphorylation site that regulates the conformation GlaxoSmithKline compound GSK1838705, which is an of the activation loop (Fig. 1). Full-length endogenous ATP-competitive inhibitor of ALK, insulin-like growth ALK supports tumor formation in NBs and glioblasto- factor (IGF)-1R, and insulin receptor (35). This compound mas, where it is suggested to be activated by the ligands is of special interest because of its good pharmacokinetic pleiotropin and midkine (22, 23); however, the impor- properties and excellent oral bioavailability (see Table 1 tance of the ligands is under debate (24). In ALCL and and Fig. 2). NSCLC, ALK is activated by chromosomal rearrange- The only compound currently undergoing clinical test- ments leading to the fusion of the ALK gene to another ing is the Pfizer compound PF-02341066 (crizotinib; gene, resulting in its dimerization and persistent activa- Fig. 2), which was initially developed as an inhibitor of tion. In NB, specific activating mutations within the c-Met (36) but was found to have a similar IC50 for ALK kinase domain of ALK have been described. In all of (24 nM). In vitro, it showed antiproliferative activity in these cases, ALK inhibition may be an effective thera- ALCL cell lines (Karpas-299 and SU-DHL-1) leading to peutic strategy; however, currently no ALK inhibitors cell cycle arrest and apoptosis. The rather moderate side have been approved for clinical cancer therapy. Several effects were predicted by the fact that it could be admi- different classes of small molecules that have been iden- nistered repeatedly in mice, dogs, and primates at con- tified as ALK inhibitors were shown to have marked centrations up to 200 mg/kg/day for up to 30 days (36). It antitumor efficacy in preclinical models. In contrast, was tested in a monotherapy trial in heavily pretreated attempts to abrogate NPM-ALK expression via siRNA NSCLC patients with tumors harboring a rearrangement were not successful, which may be due to the long half- of EML4-ALK. The overall response rate was 64% and the life (>48 h) of the NPM-ALK protein (25). Piva and disease control rate reached 90%, with a median duration colleagues (26) found that antagonizing the NPM-ALK of treatment of 25.5 weeks (37). Based on these promising mRNA with small hairpin RNA through retroviral trans- results, at the end of 2009, Pfizer began to screen 1,500 fection was a more successful approach and established patients with NSCLC for ALK translocations. Eighty-two the essential function of this fusion protein in the pro- (5.5%) of these patients had an ALK translocation þ liferation of ALK cell lines in vitro as well as in a murine (although in some cases not involving EML4) and were ALCL engraftment model. therefore eligible for PF-02341066 (crizotinib) treatment. The ALK inhibitors staurosporine and 7-hydroxystaur- It is interesting to note that this patient subgroup osporine (UCN-01) are natural products that are isolated consisted mostly of relatively young nonsmokers with
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Table 1. Selected small-molecule ALK-TK inhibitors and imatinib
Inhibitor Company Chemical type Binding site IC50 (nM) Reference Clinical trial
UCN-01 Natural comp. 7-Hydroxystaurosporine ATP pocket 123–150 (74) Yes CEP-14083 Cephalon Staurosporine-derived ATP pocket 2 (29) No CEP-14513 Cephalon Staurosporine-derived ATP pocket 4 (29) No Geldanamycin Natural comp. Benzoquinone HSP90 HSP70 (31, 75) Yes Herbimycin Natural comp. Benzoquinone HSP90 HSP70 (32) No NVP-TAE684 Novartis Diaminopyrimidine ATP pocket <10 (34) No GSK1838705 GlaxoSmithKline Diaminopyrimidine ATP pocket 0.5 (35) Yes PF-02341066 Pfizer Aminopyridine-derived ATP pocket 11–24 (5) Yes (crizotinib) Compound 18 Cephalon Diaminopyrimidine ATP pocket 4 (30) No WZ-5–126 None Unknown ATP pocket 3 (76) No Imatinib Novartis Diaminopyrimidine ATP pocket — (77) Yes
NOTE: Imatinib, the BCR-ABL inhibitor and a founding member of the group of TK inhibitor drugs, has been added as reference.
wild-type EGFR and KRAS. More than 91% of the refractory HL. In 3 of 4 patients, a rapid reduction of tumor included patients had received at least 1 previous ther- mass was observed; however, the clinical responses were apy, and, remarkably, 41% had received 3 or more. transient (41). In patients with HL and NHL, investigators Clinical activity was reported for 90% of the patients attempted to bind CD30 to the toxin ricin or 131I. However, [57% responses (1 complete and 46 partial) and 33% stable the presence of side effects such as decreases in serum disease] during the mean treatment duration of 6.4 albumin, edema, and hypotension with the ricin, or severe months. Only grade 1 and 2 gastrointestinal side effects hematotoxicity in the case of the Iodine-131-coupled anti- were reported (38). This result is encouraging, consider- body, dampened the enthusiasm for these types of treat- ing the generally poor prognosis for this class of patients. ments (42–44). In a seminal phase 1/2 study by Ansell and Crizotinib belongs to the TK inhibitor family, the most colleagues (45), it was shown that a fully human anti- prominent member of which is imatinib (Gleevec; Novar- CD30 immunoglobulin G1 monoclonal antibody (MDX- tis). Based on the experiences with this substance, we 060) was well tolerated at concentrations up to 15 mg/kg know that resistance mutations can eventually occur after in a mixed patient population, including patients with HL prolonged treatment. This underscores the need for a and ALCL. Despite high expectations, however, only 6 of wide range of chemically different ALK inhibitors. 72 patients showed a clinical response. In a more recent Indeed, such resistance mutations in the kinase domain study with 39 heavily pretreated HL patients (82% of of ALK (C1156Y and L1196M) were recently observed to whom had failed at least one prior therapy regimen) develop in a patient, and their functional importance was and 41 systemic ALCL patients using the anti-CD30 anti- confirmed by in vitro experiments (39). In addition, the body SGN-30 (Seattle Genetics), none of the HL patients first impressive clinical results regarding the use of cri- had an objective response (46). In contrast, in the ALCL zotinib in patients with inflammatory myofibroblastic arm, 2 patients achieved a complete response (4.9%) and 5 tumor (IMT), about half of whom carry the Ran-binding showed a partial response (12%), pointing to a limited protein 2-ALK (RanBP2-ALK) translocation, were clinical activity of this antibody. Side effects were rather recently presented (40). mild, with fatigue and nausea being the most frequently reported symptoms. In a phase 2 study in which the same CD30 Antibodies antibody was tested in CD30-positive, heavily pretreated patients with cutaneous ALCL, lymphomatoid papulosis, CD30 is a trans-membrane protein that belongs to the and transformed mycosis fungoides, SGN-30 showed tumor necrosis factor receptor family. It is expressed on clinical activity (complete or partial response) in 16 out activated (but not resting) B and T cells, and in addition to of 23 patients. Specifically, in cutaneous ALCL, clinical its expression on ALCL cells, it can also be found on Reed- activity was seen in all patients, with responses in 82% of Sternberg cells in Hodgkin’s lymphoma (HL). CD30 anti- the treated patients (47). These are encouraging results, bodies and derivatives were tested in patients with this especially given the mild side-effects profile. However, disease >15 years ago. In one of the first promising studies, these data have to be considered in context with current the anti-CD30 monoclonal antibody was bound to treatment strategies using pegylated liposomal doxoru- saporin, a protein with ribosome-inactivating activity. bicin and gemcitabine, which have response rates for This immunotoxin was given to 4 patients with pretreated, cutaneous ALCL ranging from 69% to 80% (48). The latest
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Novel Drugs for ALCL
H CI N HO O N
O HN N NH S O
N N O O 7-Hydroxystaurosporine H C H 3 (UCN-01) N
H3CO NVP-TAE684 NHCH3
N H O N
H Me N N N N N O N N NH O H nPr CEP-14083 O F NH N N N H N N O O H NH GSK1838705 N O N NMe Figure 2. Organic structures of 2 N N NH selected small-molecule ALK-TK O 2 CI N inhibitors and imatinib. The CEP-14513 F – structure of the substance WZ-5 i-Bu O 126 is not given because it was N N not publicly available at the time of O CI submission. O O PF-02341066 (Crizotinib) N N H H O O OH O O Cl N O N
(Me)2NC(O)CH2 N NH N NH OCONH2 O Geldanamycin Compound 18
O
N O O N NHN N H O CH CH3 3 O O HN
O O O N O N Herbimycin NH2 Imatinib
addition to these CD-30 antibody-based treatment strate- patients (86%) tumor regression was observed, and þ gies is SGN-35 (brentuximab vedotin; Seattle Genetics). In among the 11 complete remissions, 2 systemic ALK this case, the CD-30 antibody is coupled to the antitubulin ALCL cases were found (49). The success of this coupled agent monomethyl auristatin E (MMAE) with an enzyme- antibody is even more surprising in light of the limited cleavable dipeptide linker. This drug has been tested in 42 activity of the CD30 antibody alone (SGN-30) in systemic patients with CD30-positive relapsed or refractory HL ALCL, and stresses the role of the attached cytoskeleton (n ¼ 40) or ALCL (n ¼ 2) patients with a median of 3 toxin. Another appealing feature of the armed CD-30 previous therapy regimens. It is remarkable that in 36 of 42 antibody strategy is that, in contrast to ALK inhibitors,
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