{"doi":"10.1002/ajh.26084","title":"Targeting constitutively active <scp>STAT3</scp> in chronic lymphocytic leukemia: A clinical trial of the <scp>STAT3</scp> inhibitor pyrimethamine with pharmacodynamic analyses","abstract":"To the Editor: Despite the recent development of targeted therapies, chronic lymphocytic leukemia (CLL) remains incurable. Survival of CLL cells depends on constitutively activated signaling pathways that converge on a small number of transcription factors which mediate the altered gene expression that underlies the pathobiology of CLL. One such oncogenic transcription factor, which is downstream of both B cell receptor signaling and cytokines that drive B cell proliferation and survival, is STAT3. STAT3 regulates the expression of genes controlling central cellular events, including proliferation, survival, and pluripotency. In essentially all patients with CLL, STAT3 is phosphorylated on serine-727,1 which drives changes in gene expression underlying the pathogenesis of this disease.2 Through a chemical biology approach, we identified the anti-microbial agent pyrimethamine as an inhibitor of STAT3 transcriptional function.3, 4 To test the hypothesis that an inhibitor of the transcriptional function of STAT3 would have therapeutic benefit in CLL, we first evaluated the effects of pyrimethamine on CLL cells in vitro (methods found in supplementary material). Pyrimethamine caused a dramatic decrease in viable CLL cells, and did so through the induction of apoptosis (Figure S1). Peripheral blood mononuclear cells (PBMC) from healthy donors showed little effect from pyrimethamine, consistent with the known excellent safety profile of this drug. To identify genes regulated by STAT3 in CLL cells, which could serve as biomarkers for STAT3 inhibition, we first started with a set of 361 genes known to be upregulated in CLL cells compared to normal B lymphocytes5 (Figure S2A). We then filtered these genes based on regulation by STAT3 in independent data sets, or STAT3 binding in proximity to the gene by chromatin immunoprecipitation (ChIP). From this analysis, we identified five genes (AIM2, ATXN1, ENPP2, GAB1, and ID3) that showed increased expression in CLL cells compared to healthy B lymphocytes, and which had the criteria of direct STAT3 target genes. When primary CLL cells were treated ex vivo with pyrimethamine, decreased expression of all five STAT3 signature genes was consistently observed (Figure S2B). As expected, lymphocytes purified from the blood of healthy donors showed minimal expression of these genes and no significant change with pyrimethamine treatment. Given that pyrimethamine decreased the expression of STAT3 target genes and the survival of CLL cells in vitro, along with its known excellent safety profile, we designed a clinical trial to assess the efficacy of pyrimethamine as a single agent in patients with relapsed refractory CLL. This trial was initiated prior to the introduction of BTK, PI3K, or BCL-2 inhibitors into clinical use for this disease. Sixteen heavily pretreated patients, with a median of six prior therapies, enrolled on the phase one portion of this study. Patient characteristics are provided in Table S1. Three patients each were enrolled on cohorts one (12.5 mg daily) and two (25 mg daily) with no dose-limiting toxicities (DLTs) and no significant drug-related toxicities (Table S2). Cohort three (50 mg daily) enrolled 10 patients, also with no DLTs observed. The maximum tolerated dose was not reached at doses up to 50 mg daily. Plasma levels of pyrimethamine increased progressively in samples obtained during the first 2 weeks of treatment, with apparent steady state conditions achieved after dosing for 2 weeks. The steady state plasma concentration of pyrimethamine increased linearly with escalation of the daily dose from 12.5 to 50 mg (Figure S3 and Table S3). The geometric mean steady state concentration of pyrimethamine in plasma was 6.17 μM for the five patients with evaluable samples who received the 50 mg daily dose, somewhat less than the target concentration of 10 μM projected for maximal STAT3 inhibition.3, 4 The steady state concentration of pyrimethamine in PBMCs, was also linearly rel","journal":"American Journal of Hematology","year":2020,"id":100399,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":29,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9539,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2020-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":98179,"name":"Sarah R. Walker","orcid":"0000-0001-6465-241X","position":1,"is_corresponding":false},{"id":493334,"name":"Lisa N. Heppler","orcid":null,"position":2,"is_corresponding":false},{"id":66056,"name":"Svitlana Tyekucheva","orcid":"0000-0002-3119-6507","position":3,"is_corresponding":false},{"id":493335,"name":"Erik A. Nelson","orcid":null,"position":4,"is_corresponding":false},{"id":492731,"name":"Josephine L. Klitgaard","orcid":"0000-0002-4441-2643","position":5,"is_corresponding":false},{"id":493336,"name":"Maria Nicolais","orcid":null,"position":6,"is_corresponding":false},{"id":493337,"name":"Yasmin Kroll","orcid":null,"position":7,"is_corresponding":false},{"id":252122,"name":"Michael Xiang","orcid":"0000-0003-0170-9321","position":8,"is_corresponding":false},{"id":441453,"name":"Jennifer Yeh","orcid":"0000-0003-3144-6702","position":9,"is_corresponding":false},{"id":493338,"name":"M. Chaudhury","orcid":null,"position":10,"is_corresponding":false},{"id":493339,"name":"Zachary T. Giaccone","orcid":null,"position":11,"is_corresponding":false},{"id":3260,"name":"Stacey M. Fernandes","orcid":"0000-0002-5061-5199","position":12,"is_corresponding":false},{"id":11124,"name":"Eric D. Jacobsen","orcid":"0000-0003-3014-0023","position":13,"is_corresponding":false},{"id":343821,"name":"David C. Fisher","orcid":"0000-0001-6915-8584","position":14,"is_corresponding":false},{"id":3267,"name":"Arnold S. Freedman","orcid":"0000-0002-8227-8916","position":15,"is_corresponding":false},{"id":233541,"name":"Matthew S. Davids","orcid":"0000-0003-4529-2003","position":16,"is_corresponding":false},{"id":492732,"name":"Jeffrey G. Supko","orcid":"0000-0002-4599-3267","position":17,"is_corresponding":false},{"id":3263,"name":"Catherine J. Wu","orcid":"0000-0002-3348-5054","position":18,"is_corresponding":false},{"id":398018,"name":"David A. Frank","orcid":"0000-0002-7698-8364","position":19,"is_corresponding":false},{"id":3252,"name":"Jennifer R. Brown","orcid":"0000-0003-2040-4961","position":0,"is_corresponding":true}],"reference_count":7,"raw_metadata":null,"created_at":"2026-07-18T22:38:35.493878Z","pmid":"33373063","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}