A Novel Target Antigen for the Treatment of Acute Myeloid Leukemia by CAR T Cells
Authors: Paul A. Beavis, Kevin Sek, Phillip K. Darcy
Journal: Molecular Therapy (2017)

There is strong evidence that the presence of immune infiltrate in tumors is correlated with a good prognosis following therapy.1Loi S. Sirtaine N. Piette F. Salgado R. Viale G. Van Eenoo F. Rouas G. Francis P. Crown J.P. Hitre E. et al.Prognostic and predictive value of tumor-infiltrating lymphocytes in a phase III randomized adjuvant breast cancer trial in node-positive breast cancer comparing the addition of docetaxel to doxorubicin with doxorubicin-based chemotherapy: BIG 02-98.J. Clin. Oncol. 2013; 31: 860-867Crossref PubMed Scopus (1133) Google Scholar, 2Galon J. Costes A. Sanchez-Cabo F. Kirilovsky A. Mlecnik B. Lagorce-Pagès C. Tosolini M. Camus M. Berger A. Wind P. et al.Type, density, and location of immune cells within human colorectal tumors predict clinical outcome.Science. 2006; 313: 1960-1964Crossref PubMed Scopus (4564) Google Scholar This has led to the development of T cell-based immunotherapies for cancer. One such therapy has involved isolating and expanding tumor reactive lymphocytes from the tumor, called tumor-infiltrating lymphocyte (TIL) therapy. This form of adoptive immunotherapy utilizing TILs has shown significant success in patients with advanced melanoma, with greater than 50% of patients responding to treatment.3Dudley M.E. Wunderlich J.R. Yang J.C. Sherry R.M. Topalian S.L. Restifo N.P. Royal R.E. Kammula U. White D.E. Mavroukakis S.A. et al.Adoptive cell transfer therapy following non-myeloablative but lymphodepleting chemotherapy for the treatment of patients with refractory metastatic melanoma.J. Clin. Oncol. 2005; 23: 2346-2357Crossref PubMed Scopus (1363) Google Scholar However, the broad use of TIL therapy has been precluded by the inability to isolate sufficient number of tumor reactive T cells from most cancers. To overcome this problem, the genetic modification of T cells with αβ T cell receptor (TCR) transgenes4Parkhurst M.R. Joo J. Riley J.P. Yu Z. Li Y. Robbins P.F. Rosenberg S.A. Characterization of genetically modified T-cell receptors that recognize the CEA:691-699 peptide in the context of HLA-A2.1 on human colorectal cancer cells.Clin. Cancer Res. 2009; 15: 169-180Crossref PubMed Scopus (95) Google Scholar or recombinant receptors, called chimeric antigen receptors (CARs),5Grupp S.A. Kalos M. Barrett D. Aplenc R. Porter D.L. Rheingold S.R. Teachey D.T. Chew A. Hauck B. Wright J.F. et al.Chimeric antigen receptor-modified T cells for acute lymphoid leukemia.N. Engl. J. Med. 2013; 368: 1509-1518Crossref PubMed Scopus (2489) Google Scholar have been developed that arm the T cell with anti-tumor activity regardless of major histocompatibility I (MHCI) expression. The promise of this approach has been recently highlighted in certain B cell malignancies, such as acute lymphoid leukemia (ALL) and chronic lymphoid leukemia utilizing CAR T cells specific for CD19, and has resulted in objective responses in up to 90% of ALL patients.6Maude S.L. Frey N. Shaw P.A. Aplenc R. Barrett D.M. Bunin N.J. Chew A. Gonzalez V.E. Zheng Z. Lacey S.F. et al.Chimeric antigen receptor T cells for sustained remissions in leukemia.N. Engl. J. Med. 2014; 371: 1507-1517Crossref PubMed Scopus (3540) Google Scholar These striking results have raised the prospect of utilizing CAR T cells targeting novel antigens for other malignancies, including acute myeloid leukemia (AML). Previous CAR T cell therapy trials in AML did not result in long term responses, and CAR T cells against AML antigens, such as CD123 and CD33, exhibited unwanted toxicity toward normal hematopoietic progenitor cells (HPCs). Therefore, there is interest in generating CAR T cells with higher specificity toward an antigen expressed exclusively on AML cells that spare normal myeloid progenitor cells. In this issue of Molecular Therapy, Tashiro et al.7Tashiro H. Sauer T. Shum T. Parikh K. Mamonkin M. Omer B. Rouce R.H. Lulla P. Rooney C.M. Gottschalk S. Brenner M.K. Treatment of acute myeloid leukemia with T cells expressing chimeric antigen receptors directed to C-type lectin-like molecule 1.Mol. Ther. 2017; 25 (this issue): 2202-2213Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar investigate the effectiveness of CAR T cells targeting the C-type lectin-like molecule 1 (CLL-1). This membrane glycoprotein serves as a potential target antigen given its high expression on a majority of AML cells (expressed in 18/19 patients tested) and, importantly, its absence from normal hematopoietic stem cells. CLL-1 has previously been validated as a target in AML through the use of monoclonal antibodies.8Zhao X. Singh S. Pardoux C. Zhao J. Hsi E.D. Abo A. Korver W. Targeting C-type lectin-like molecule-1 for antibody-mediated immunotherapy in acute myeloid leukemia.Haematologica. 2010; 95: 71-78Crossref PubMed Scopus (75) Google Scholar In the new study, the authors first demonstrate antigen-specific function of T cells that had been gene-modified with a second-generation CAR receptor (anti-CLL-1BBζ). The data included cytotoxicity, proliferation, and cytokine release against both CLL-1+ AML lines and primary AML patient samples. Furthermore, anti-leukemic activity of adoptively transferred anti-CLL-1 T cells against disseminated AML in a xenograft mouse model was shown. An important finding was that anti-CLL-1BBζ T cells were found to be cytotoxic against mature monocytes, but not to normal myeloid progenitor cells, in vitro. This feature should allow full myeloid recovery following elective ablation of anti-CLL-1 T cells, thereby protecting the patient from any dangerous and long term toxic side effects. There have been several reports of tumor relapse in ALL patients following CAR 19 T cell therapy as a result of the emergence of CD19− variant clones.9Gardner R. Wu D. Cherian S. Fang M. Hanafi L.A. Finney O. Smithers H. Jensen M.C. Riddell S.R. Maloney D.G. Turtle C.J. Acquisition of a CD19-negative myeloid phenotype allows immune escape of MLL-rearranged B-ALL from CD19 CAR-T-cell therapy.Blood. 2016; 127: 2406-2410Crossref PubMed Scopus (472) Google Scholar One potential approach to circumvent this problem for the treatment of AML and other cancers may be to redirect T cells to multiple antigens. For example, genetic modification of T cells with CARs recognizing both CLL-1 and Lewis Y for AML may prove to be more effective long term and protect against potential editing of tumor antigen and the emergence of antigen-negative clones. Given that Tashiro et al.7Tashiro H. Sauer T. Shum T. Parikh K. Mamonkin M. Omer B. Rouce R.H. Lulla P. Rooney C.M. Gottschalk S. Brenner M.K. Treatment of acute myeloid leukemia with T cells expressing chimeric antigen receptors directed to C-type lectin-like molecule 1.Mol. Ther. 2017; 25 (this issue): 2202-2213Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar found CLL-1 expression to be fairly heterogenous (33%–99% expression), targeting multiple tumor antigens would also likely enhance efficacy in patients with variable expression of CLL-1. Another issue for effective CAR T cell therapy is the immunosuppressive tumor microenvironment. Tumors can evade immune recognition by activating various checkpoint pathways, including PD-1 and CTLA-4. Tumors often show increased expression of ligands for these receptors, which results in a significant reduction of effector cell function. In the clinic, immunotherapy mediated by antibodies such as pembrolizumab, which blocks interaction of PD-1 with PD-L1 or PD-L2, or ipilimumab, which blocks the interaction between CTLA-4 and CD80/CD86, has resulted in striking effects in cancers such as melanoma, lung and renal cancers, and, in some cases, is durable. There have been several reports in preclinical mouse models demonstrating that checkpoint blockade using anti-PD-1 can significantly augment CAR T cell responses.10John L.B. Devaud C. Duong C.P. Yong C.S. Beavis P.A. Haynes N.M. Chow M.T. Smyth M.J. Kershaw M.H. Darcy P.K. Anti-PD-1 antibody therapy potently enhances the eradication of established tumors by gene-modified T cells.Clin. Cancer Res. 2013; 19: 5636-5646Crossref PubMed Scopus (523) Google Scholar, 11Beavis P.A. Henderson M.A. Giuffrida L. Mills J.K. Sek K. Cross R.S. Davenport A.J. John L.B. Mardiana S. Slaney C.Y. et al.Targeting the adenosine 2A receptor enhances chimeric antigen receptor T cell efficacy.J. Clin. Invest. 2017; 127: 929-941Crossref PubMed Scopus (200) Google Scholar, 12Cherkassky L. Morello A. Villena-Vargas J. Feng Y. Dimitrov D.S. Jones D.R. Sadelain M. Adusumilli P.S. Human CAR T cells with cell-intrinsic PD-1 checkpoint blockade resist tumor-mediated inhibition.J. Clin. Invest. 2016; 126: 3130-3144Crossref PubMed Scopus (586) Google Scholar Based on these reports, several clinical trials are currently underway examining adoptively transferred T cells in combination with checkpoint blockade (clinicaltrials.gov: NCT02652455). A recent case study report described a patient with diffuse large B cell lymphoma who had progressive disease following CAR 19 T cell therapy but responded after administration of pembrolizmumab. This result correlated with expansion of CAR 19 T cells and an increase in transgene copy number.13Chong E.A. Melenhorst J.J. Lacey S.F. Ambrose D.E. Gonzalez V. Levine B.L. June C.H. Schuster S.J. PD-1 blockade modulates chimeric antigen receptor (CAR)-modified T cells: refueling the CAR.Blood. 2017; 129: 1039-1041Crossref PubMed Scopus (322) Google Scholar This observation clearly warrants testing of CAR T cell combination therapy with checkpoint blockade in other blood cancers, such as AML, and also in solid tumors. The relative effect of targeting other immunosuppressive pathways could further improve CAR T cell activity. A recent study reported that targeting the CD73/adenosine pathway using antagonist drugs blocking the A2A adenosine receptor could augment CAR T cell activity against several syngeneic Her-2+ tumors in mice.11Beavis P.A. Henderson M.A. Giuffrida L. Mills J.K. Sek K. Cross R.S. Davenport A.J. John L.B. Mardiana S. Slaney C.Y. et al.Targeting the adenosine 2A receptor enhances chimeric antigen receptor T cell efficacy.J. Clin. Invest. 2017; 127: 929-941Crossref PubMed Scopus (200) Google Scholar Thus, combination approaches that target the immunosuppressive effects of adenosine and other checkpoint pathways may result in a greater proportion of patients responding to CAR T cell adoptive immunotherapy. A serious concern with immunotherapies such as CAR T cells is the potential for on-target, off-tumor activity leading to fatal toxicity. This has been reported in several trials including patients with CAR19 T cells14Morgan R.A. Yang J.C. Kitano M. Dudley M.E. Laurencot C.M. Rosenberg S.A. Case report of a serious adverse event following the administration of T cells transduced with a chimeric antigen receptor recognizing ERBB2.Mol. Ther. 2010; 18: 843-851Abstract Full Text Full Text PDF PubMed Scopus (1742) Google Scholar (CAR19 trial; clinicaltrials.gov: NCT02535364). Thus, targeting an antigen on tumor cells that spare normal hematopoietic cells would be most favorable. In the current study by Tashiro et al.,7Tashiro H. Sauer T. Shum T. Parikh K. Mamonkin M. Omer B. Rouce R.H. Lulla P. Rooney C.M. Gottschalk S. Brenner M.K. Treatment of acute myeloid leukemia with T cells expressing chimeric antigen receptors directed to C-type lectin-like molecule 1.Mol. Ther. 2017; 25 (this issue): 2202-2213Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar targeting the CLL-1 antigen appears to be ideal, given it is expressed predominately on AML cells, and, although it is present on mature monocytes, it is not expressed on immature myeloid precursor cells. This should allow reconstitution of the myeloid population using methods that enable elective ablation of the CAR T cells. One potential approach, as shown in the current study, utilizes CAR T cells expressing an inducible caspase-9 suicide gene that can be effectively eliminated by administration of the activating dimerizer drug (chemical inducer of dimerization [CID]). Importantly, this approach has been validated in the clinic.15Di Stasi A. Tey S.K. Dotti G. Fujita Y. Kennedy-Nasser A. Martinez C. Straathof K. Liu E. Durett A.G. Grilley B. et al.Inducible apoptosis as a safety switch for adoptive cell therapy.N. Engl. J. Med. 2011; 365: 1673-1683Crossref PubMed Scopus (1040) Google Scholar The use of CAR T cells engineered with a suicide gene therefore provides an inbuilt safety mechanism for patients. It will be important for future studies to investigate the toxicity associated with anti-CLL-1 CAR T cells in an immunocompetent setting, potentially in a self- antigen model, and whether deletion of CLL-1-specific CAR T cells using this system increases the risk of relapse. In summary, the era of immunotherapy has entered an exciting new phase with recent results in patients using checkpoint inhibitors for various cancers or CAR T cells directed against CD19 for B cell malignancies. In the future, targeting of T cells using CARs directed against other antigens, such as CLL-1, that are predominately expressed on tumor cells and not on normal hematopoietic stem cells may significantly broaden both the effectiveness and safety of adoptive T cell immunotherapy for many cancers, including AML. P.A.B. is supported by a National Breast Cancer Foundation Fellowship (ID# ECF-17-005). P.K.D. is supported by an NHMRC Senior Research Fellowship (APP1041828). Treatment of Acute Myeloid Leukemia with T Cells Expressing Chimeric Antigen Receptors Directed to C-type Lectin-like Molecule 1Tashiro et al.Molecular TherapyJune 5, 2017In BriefTashiro et al. demonstrate that T cells expressing a C-type lectin like molecule-1-specific chimeric antigen receptor (CLL-1 CAR) exert potent anti-leukemic activity against CLL-1+ AML cell lines as well as AML patient samples. Normal hematopoietic stem cells (HSCs) do not express CLL-1 and are spared by this CAR-T treatment. Full-Text PDF Open Archive