Trametinib for the Treatment of IGHV4–34, MAP2K1 Mutant Variant Hairy Cell Leukemia

Variant hairy cell leukemia (vHCL) is an uncommon disorder accounting for 10% of all HCL cases [1]. The difference in therapeutic response to chemotherapy between patients with classic HCL (cHCL) and vHCL is impressive; whereas up to 90% of cHCL patients achieve a complete remission (CR) with purine analog therapy alone, fewer than 50% of variant patients do. In 2011, BRAF mutations, which activate the RAF/MEK/ERK pathway, were identified in cHCL [2]. In contrast, BRAF is not mutated in vHCL. Consequently, vHCL is no longer considered biologically related to cHCL and is included among the unclassifiable splenic B-cell leukemia/lymphomas in the WHO classification. Variant HCL is characterized by frequent IGHV4–34 usage, lack of CD25 positivity and mutations or deletions of TP53 [3]. IGHV4–34 (+) vHCL has a worse prognosis compared to classic HCL and IGHV4–34(−) vHCL [4], with inferior responses to chemotherapy and shorter durations of remission. Recently, activating mutations of MAP2K1, which encodes MEK1, have been identified in up to half of vHCL cases [5]. These mutations result in activation of MEK/ERK signaling. Trametinib is a MEK inhibitor which is FDA approved for the treatment of patients with BRAF p.V600E mutant melanoma. This agent reversibly binds to MEK1 and MEK2, preventing downstream phosphorylation of ERK and decreasing cellular proliferation and survival. We hypothesized that trametinib would have activity in MAP2K1-mutant vHCL.

The patient is a 52-year-old man who initially presented with splenomegaly and lymphocytosis and was diagnosed with BRAF wild type, variably positive CD25, IGHV4–34(+) vHCL in 2005. IGHV mutational analysis was performed on PBMCs (81% hairy cells on manual differential) collected in 2013. Briefly, Sanger sequencing was performed on cDNA generated via RNA extraction, reverse transcription and PCR amplification of the entire IGHV region using VH-CH and VH-JH primer sets. This yielded an amplicon with 100% identity to the reference IGHV4–34 gene according to IgBlast. His previous treatments included cladribine, BL22, pentostatin/rituximab, splenectomy, single-agent rituximab, ibrutinib, bendamustine/rituximab and allogeneic transplantation from a matched unrelated donor (summarized in Table 1). Of note, during his disease course, serum CD25, while not typically a biomarker in vHCL patients, was consistently elevated when his leukemia was active. CD25 expression by flow cytometry ranged from 10% to 30% to nearly 100% of the hairy cell population, with the lower expression occurring after rituximab or ibrutinib treatment. The patient experienced disease relapse day +350 post-transplant when he developed skin nodules and generalized skin rash (Figure 1(A)). The rash appeared clinically consistent with acute GVHD. However, biopsies of both the nodules and rash showed cutaneous relapse of vHCL (Figure 1(C)). In addition, serum CD25 was rising, peaking at 29,219 U/mL (Figure 1(G)). He had leukemic lymphocytosis (WBC 16,000/μL, absolute lymphocyte count 6080/μL) but no cytopenias or lymphadenopathy. Bone marrow biopsy showed vHCL involvement comprising 40% of the marrow cellularity with strong expression of CD25. He was consented for paired next-generation sequencing of blood and skin biopsy with the Ilumina TruSight Tumor 26 panel, a commercially available gene amplicon capture panel performed on the Ilumina MiSeq platform. This revealed a somatic MAP2K1 p.K57N mutation (VAF 43.26% in skin and 20.08% in blood) that has been shown to constitutively activate MEK [5]. No other somatic variants were detected. Given disease involvement in both skin and blood, it was not possible to perform true germline sequencing. However, the difference in mutation VAF between skin and blood was consistent with an acquired somatic mutation.

Table 1.

Timeline

2005	Presented with symptomatic splenomegaly and diagnosed with vHCL on basis of variable CD25 expression
2007	Cladribine treatment
2007	Rapid regrowth of spleen
2007	BL22 study- no response, developed antibody to immunotoxin
2007	Pentostatin/rituximab, no response
2007	Splenectomy, with clinical improvement
2013	Lymphocytosis, thrombocytopenia, anemia, BRAFV600E negative
2013	Ibrutinib on study with disease progression
2014	Bendamustine and rituximab with anaphylactoid reaction to rituximab
2014	Continued with single-agent bendamustine achieving stable disease.
April 2015	Allogeneic transplant: matched unrelated donor PBSCT using RIC (fludarabine/busulfan/ATG)
May 2015	GVHD of the skin which was steroid responsive
February 2016	Bone marrow biopsy: 20% cellularity, immunohistochemical staining for Pax5 demonstrates the hairy cell leukemia to account for 40% of the marrow cellularity
March 2016	Biopsies of rash and nodules behind both ears show leukemic cell involvement and were negative for GVHD.
February 2016	NGS performed on skin biopsy showed MAP2K1 K57N mutation.
March 2016	Continued erythematous maculopapular rash over back, shoulders and thighs
May 2016	Trametinib treatment start date
May 2016	Pustular acneiform rash over sun exposed areas (face, neck and to lesser extent bilateral forearms).
June 2016	Bone marrow biopsy: 40% cellular bone marrow with persistent involvement by hairy cell leukemia with variant immunophenotype and partially preserved trilineage hematopoietic reserves. 60% CD20-positive cells
August 2016	Skin nodules have almost completely resolved. Cutaneous toxicities are improved except when he experiences excess sun exposure.
October 2016	Skin biopsy: No evidence of cutaneous involvement by hairy cell leukemia cells. Grover’s Disease- rash produced by Trametinib
January 2017	Bone marrow biopsy: 70% cellularity bone marrow with hairy cell leukemia accounting for 40% of the marrow cellularity as noted by staining for Pax5 and CD20.
January 2017	CT neck/chest/abdomen/pelvis: Interval increase in size of the lymph nodes in the neck. Index cervical node measurements include 6 × 7 mm → 11 × 10 mm, 7 × 6 mm → 11 × 10 mm and 8 × 6 mm → 12⋅×⋅10 mm. No new or progressive lymphadenopathy in chest, abdomen or pelvis
February 2017	Skin biopsy: Subacute spongiotic dermatitis, with focal interface reaction. Diagnostic evidence of leukemia cutis is not identified

Figure 1.

(A) Leukemia cutis rash and postauricular nodules before trametinib treatment and (B) after 2 months of trametinib therapy. H&E staining and immunohistochemistry of phospho–ERK on skin biopsy pre-(C,D) and post-treatment (E,F). Scale bar = 500 µm. (G) Serum IL-2 receptor (CD25) before and after trametinib treatment. Treatment start is indicated by vertical dashed line.

As the patient had exhausted the majority of available treatment options, he was prescribed trametinib 2 mg PO daily, according to approved melanoma dosing. Within a week of therapy initiation, his skin nodules were markedly diminished in size and his generalized rash had resolved (Figure 1(B)). We also noted a significant decrease in serum CD25 soon after starting trametinib (Figure 1(G)). He did develop a new acneiform facial rash consistent with drug toxicity. This improved with oral doxycycline and sun avoidance and did not require a trametinib dose reduction. We observed near complete resolution of skin nodules behind each ear and disappearance of visible skin rash following cycle 2 of therapy. His WBC and ALC decreased to the reference range (WBC 6800/μL, absolute lymphocyte count 2520/μL) by the end of cycle 3. Repeat bone marrow biopsy showed unchanged hairy cell index. Skin biopsies were repeated and phosphoERK staining of skin biopsies pre- and post-trametinib was performed (Figure 1(C–F)). This showed diminished lymphocyte involvement on H&E staining (Figure 1(C,E)) with a decrease in phosphoERK immunostaining (Figure 1(D,F)), indicative of decreased signaling downstream of MEK and consistent with on target trametinib effects. The patient remained on trametinib with resolution of rash; however, restaging analysis at completion of cycle 6 showed stable disease, with bone marrow biopsy showing unchanged hairy cell index, and CT scans of the neck, chest, abdomen and pelvis showing no thoracic or abdominopelvic lymphadenopathy, but a slight increase in small neck adenopathy not meeting criteria for progression. During this time period, serum CD25 decreased to minimal elevation (1709 U/mL, ULN 1100 U/mL), consistent with decreased disease activity.

Variant HCL-associated MAP2K1 mutations are found within sequences encoding the negative regulatory region domain or the kinase catalytic domain [5]. Binding of allosteric MEK inhibitors to MEK1, and thus, drug sensitivity requires hydrophobic interactions with multiple residues in the activation loop and alpha C helix, resulting in stabilization of an inactive catalytic site conformation [6]. Our patient’s K57N mutation does not disrupt drug binding by the MEK inhibitor selumetinib [7], which has the same binding site as that of trametinib and other allosteric MEK inhibitors [8]. The binding of K57N-mutant MEK1 to trametinib has not been experimentally verified, though the binding site of all studied allosteric MEKi is similar [9]. Some mutations, including Q56P, I103N and C121S, simultaneously result in both MEK activation and disruption of allosteric MEK inhibitor binding via alteration of amino acids important for binding [10]. These resistant mutations may occur de novo or following MEK inhibitor therapy. Notably, C121S is the most common MAP2K1 alteration in vHCL, followed by K57 mutations [5]. This raises the possibility that a subset of MAP2K1-mutated vHCL patients may not respond to MEK inhibition or may require higher doses, depending on their specific MAP2K1 mutation. However, some mutations may be context dependent. For example, the Q56P mutation has been reported to be resistant to trametinib when present in BRAF V600E + melanoma [10] and sensitive to MEK inhibition in the BRAF WT lung adenocarcinoma cell line NCI-H1437 [11]. In addition, clinical responses may occur in patients with mutations deemed ‘resistant’ in laboratory studies. Diamond et al reported a response to cobimetinib in a heavily pretreated MAP2K1 p.Q56P-mutant Erdheim-Chester disease patient [12]. We submit that restricting enrollment of MAP2K1-mutated vHCL patients to clinical trials based upon specific mutations is premature, though correlation of MAP2K1 mutations with MEK inhibitor response remains an important research question.

In metastatic melanoma patients receiving single-agent trametinib, duration of response is typically less than six months [13], so our patient’s modest response after 6 months of treatment is therefore not unexpected. He remains on trametinib due to the sustained improvement in his cutaneous leukemia. In melanoma patients who have progressed on trametinib, discontinuation may result in a rapid increase in the rate of progression, suggesting that even resistant tumors retain partial sensitivity that is clinically meaningful [13]. This phenomenon has an obvious parallel to the rapid acceleration seen in CLL patients who discontinue ibrutinib following disease progression [14]. We suggest vHCL patients taking a MEK inhibitor on study should be closely monitored during discontinuation. In addition, studies of MEKi in vHCL should include repeat MAP2K1 sequencing for patients who progress in order to identify possible additional resistance mechanisms. These investigations may provide clues to develop effective targeted combination strategies to circumvent the development of resistant disease.

To our knowledge, this is the first report of treatment of a MAP2K1-mutant vHCL patient treated with the oral MEK inhibitor trametinib. Therapy has been well tolerated, with the only adverse event being a transient acneiform rash, a known on-target toxicity, which did not require dose reduction. Thus far, he has no evidence of the most serious potential adverse events associated with trametinib, such as cardiomyopathy, ocular toxicity, cutaneous malignancy or serious skin toxicity. Dosing was based largely on the experience using trametinib for malignant melanoma. The optimal dose and duration of therapy should be explored in prospective clinical trials. The requirement for different doses of a targeted inhibitor for different malignancies is not unprecedented; for example, ibrutinib is dosed at 560 mg for mantle cell lymphoma as opposed to the 420 mg dose used in the treatment of chronic lymphocytic leukemia. In addition, variable doses and schedules of vemurafenib have been found to be effective in obtaining a response in cHCL [15,16]. Furthermore, in the initial phase-1 study of trametinib, daily doses up to 3 mg were tolerated without serious adverse events [17], suggesting that dose escalation can be explored in the context of a controlled trial. Despite the presence of MAK2K1 mutations in other B-cell malignancies, there are no published cases of other lymphomas, such as pediatric-type nodal follicular lymphoma, splenic marginal zone lymphoma or splenic diffuse red pulp small B-cell lymphoma being successfully treated with MEK inhibitors. MAP2K1 mutations are found in a subset of Langerhans histiocytosis and non-Langerhans histiocytoses, where clinical activity of trametinib has been reported in two patients [12].

In conclusion, trametinib has some single-agent activity in a variant HCL patient with a MAP2K1 p.K57N mutation and may represent a novel therapeutic approach in vHCL.