Cell Therapies Emerge for the Treatment of Acute Myeloid Leukemia


Cellular therapies are an effective option in hematologic malignancies but have been slower to develop in AML, but identifying new targets paves the way for evolving treatments.

Acute myeloid leukemia (AML) cells in blood flow - isometric view, 3D illustration: © LASZLO - stock.adobe.com

Acute myeloid leukemia (AML) cells in blood flow - isometric view, 3D illustration: © LASZLO - stock.adobe.com

Cellular therapies have shown effectiveness in treating hematologic malignancies, although their development for acute myeloid leukemia has been slower. However, identifying new targets is paving the way for advancing treatments in this area.

Conventional treatment for acute myeloid leukemia (AML) is largely dependent on hypomethylating agents (HMAs) and intensive chemotherapy. However, recent breakthroughs in cellular therapy offer promising alternatives for treating this disease.

Despite these advances, a lack of matched marrow donors, in vitro preparation, graft rejection, and relapse posttransplant present significant challenges when implementing cellular therapy treatments.

Cellular therapies like hematopoietic stem cell transplant (HSCT) and chimeric antigen receptor (CAR) T-cell therapy have warranted new research, and novel cellular therapies also show potential for treating this disease.

Challenges in AML Treatment

Several aspects make AML challenging to treat, and perhaps the primary reason is the heterogeneity of the disease.

“For FLT3-mutated and IDH1- and IDH2-mutated as well as now emerging for MLL-rearranged and PM1 with Menin inhibitors, we do have effective salvages, initially as single agents moving into combinations, showing high response rates as well as a good opportunity to get those patients for transplant with a chance at long-term survival. But for the other 50% to 60% who do not have 1 of these targetable mutations, there’s really no established therapy that has shown to be effective,” Naval G. Daver, MD, professor and director of the Leukemia Research Alliance Program in the Department of Leukemia at MD Anderson Cancer Center in Houston, Texas, said in an interview with Targeted OncologyTM.

Other challenges for treating AML include the frequent expression of targets on healthy hematopoietic progenitors. This can make targeted therapies less effective and potentially more toxic. Moreover, unique interactions in the tumor microenvironment further complicate treatment strategies.1

Daver identified a key issue as being able to move past the standard treatment combination of HMAs and venetoclax (Venclexta), which leaves room for improvement regarding survival outcomes.

“[It is] a good regimen [and] gives us good remission rates. But when you look at the long-term survival, the 3-year survival is only about 23% to 24%. So how do we make this better? Maybe by adding targeted therapies to the frontline combination, getting deeper remissions, and potentially that will improve the survival,” Daver said.

“The other question that is coming is whether we can start incorporating some forms of immunotherapy, whether these are [natural killer (NK)]-cell based [or] T-cell based, in patients who will achieve remission but often have detectable low-level disease burden,” Daver added.

CAR T-cell therapy

CAR T-cell therapy

CAR T-Cell Therapy in AML

The CAR T-cell therapies that are currently FDA-approved target CD19 or B cells in patients with B-cell acute lymphoblastic leukemia, B-cell lymphoma subtypes, mantle cell lymphoma, and multiple myeloma.2

“Currently, there is an interest in using CAR T cells for AML…But when you look at the data out there that has been published, even though there are more than 30 clinical trials that are currently open, only less than 100 patients have been reported. Most of them are in abstract form with overall response rate that is still not where we want it,” Paulina Velasquez, MD, assistant faculty member, St. Jude Children’s Research Hospital, said in an interview with Targeted OncologyTM.

However, 1 drawback of CAR T-cell therapy is that it is limited to surface cell antigens.1

“I think 1 of the reasons why this hasn't worked is there are many, many factors that can impact the effectiveness of these cells [and]…there's a lot of overlap between antigens expressed on AML blasts and on normal tissue. When you think about it, most of the antigens that we have are also expressed on normal hematopoietic progenitor cells and mature granulocytes, which is different from what we see, for example, in CD19,” Velasquez explained.

TCR Therapy in AML

T-cell receptor (TCR) therapy offers an alternative to CAR T-cell therapy that can reach intracellular targets and expand the number of targetable mutations. Additionally, TCR therapy can eliminate several antigen-presenting tumor cells.3

“On the TCR T-cell side...there are 3 different approaches. You have the naturally occurring TCR, and these are cells that are not genetically modified…Then, you have the T cells where you adoptively transfer at TCR. You can make it specific to certain AML fusions, for example. Last but not least, you can have a mix of those situations in which you are incorporating gene editing techniques to try to improve or enhance these therapies,” explained Velasquez.

Velasquez explained that the TCR-based strategy is major histocompatibility complex (MHC)-dependent, compared with CAR T cells that are MHC-independent. However, TCR therapy is limited to the HLA allele that is presenting the epitope, which restricts the number of patients that would be eligible.

Creative image of embryonic stem cells, cellular therapy: © pinkeyes - stock.adobe.com

Creative image of embryonic stem cells, cellular therapy: © pinkeyes - stock.adobe.com

CAR NK-Cell Therapy in AML

“There is continued interest in cell therapies for AML. I think we continue to work at T-cell–based strategies, but there are also other interesting approaches using other platforms like NK cells. That has been very exciting to see the data as well,” Velasquez said.

CAR-NK cell-based immunotherapy offers a beneficial tumor response-to-toxicity profile while minimizing immune-related adverse events. As of November 2023, there are 9 interventional clinical trials with 297 enrolled patients investigating CAR-NK cells in AML.Perinatal blood and stem cells appear to be more effective than adult tissue as seed cells for CAR-NK cell generation.4

Targetable Mutations in AML


When CD123 expression rises in patients with AML, overall survival (OS) and progression-free survival (PFS) decreases. CD123 expression has also been linked to a high risk of treatment failure.5

The target is more promising than CD33, as it is more highly expressed by AML cells and less expressed on healthy hematopoietic and progenitor cells. However, there is still a risk of hematopoietic toxicities with anti-CD123 agents, especially when adjuvant agents that increase CD123 expression are added to improve the targeted killing.

“…We have a CD123 CAR that is currently being tested in a clinical trial [NCT04318678],” Velasquez noted. “AML is a heterogeneous disease, and I think targeting only 1 antigen is probably not going to be the answer…We made a very specific CAR, and we tested it out. We have a panel of different construction… We did our functional characterization and collaborated with structural biology to do some protein folding prediction analysis using AlphaFold2, and then confirmed results as we went…We were able to find 1 construct that recognized both antigens.”


Almost all leukemic cells express high levels of CD33, while it has lower levels of expression on normal cells. Current research has shown that CD33 is a notable target because its loss from the surface of AML primitive cells as an antigen-negative escape is linked to worse outcomes for patients.

Off-target toxicities with anti-CD33 agents can be present, and preclinical models have identified typical hematopoietic toxicities with CD33-targeting agents, including hemocytopenia and decreased myeloid progenitor cells. However, combining CD33 CAR T-cell therapy with CD33 knockout for HSCT is 1 method that can be used to potentially reduce these toxicities.


CD7 is expressed in leukemic blasts and malignant progenitor cells in about 30% of AML cases. CD7 expression by malignant blasts is associated with chemoresistance and poor clinical outcomes.6 However, CD7 is also present in healthy cells. This means that CAR T-cell treatments targeting CD7 may attack other T and NK cells and deplete the CAR T cells after infusion.5


CD70 is another protein that is highly upregulated on AML cells, including leukemic stem cells. Two clinical trials are investigating CD70 CAR T-cells (NCT04662294) and CD70 NK cells (NCT05667155).5 Findings from a study published in Blood in 2021 reported that CD70-targeted CAR T did not affect normal hematopoiesis but required monitoring of virus-specific T-cell responses.7


Myeloid cells and AML monocytes highly express ILT3, also known as LILRB4, and ILT3 was also observed to trigger apoptosis against monocytic AML in vitro and decrease tumor burden in a xenograft model in vivo. It did not appear to cause adverse effects on normal hematopoiesis.5


Overexpression of NKG2DL is seen in solid tumors and hematologic malignancies, including AML and multiple myeloma. Tumor cells regulate NKG2DL at multiple levels, making it escape recognition by NK cells. Further evidence suggests that NKG2DL expression can serve as a predictive and prognostic marker in patients with AML.


FLT3 is 1 of the most prevalent mutations in patients with AML, making them an attractive target.

“The big change over the last couple of years has probably been the use of FLT3 inhibitors. About one-third of patients will have a FLT3 mutation, and these patients respond better and have a more durable response and improve survival with the addition of a FLT3 inhibitor to the intensive chemo backbone,” Daver said.

While FLT3 inhibitors like gilteritinib (Xospata) and midostaurin (Rydapt) can improve clinical outcomes, allogeneic HSCT is still the most effective option for FLT3-mutated AML. FLT3-targeted CAR T-cell therapy is an emerging option, and the FLT3 inhibitor crenolanib can help to upregulate FLT3-internal tandem duplication expression on the cell surface, which can improve the targeting of FLT3 CAR T.


As CLL-1 is not expressed in healthy tissues, it is a promising therapeutic target. Two trials are recruiting and investigating CLL-1-targeted CAR T-cell therapies. One early phase 1 study (NCT05252572) is evaluating the dose-limiting toxicities and incidence of treatment-emergent adverse events with anti-CLL-1 CAR T cells for the treatment of hematologic malignancies, including AML.8 The other is a phase 1/2 study (NCT04884984) of anti-CLL-1 CAR T cells for CLL-1-positive relapsed/refractory AML.

The study has a primary end point of number of adverse events and secondary end points of overall response rate, overall survival, event-free survival, and cumulative incidence of relapse.9


While CD38 is expressed in cells of hematologic malignancies and broadly expressed on immune cells, plasma cells, and erythrocytes, it is not expressed on hematopoietic stem cells. Daratumumab (Darzalex) is an anti-CD38 monoclonal antibody that has shown promise for the treatment of multiple myeloma. Up to 83% of AML cells express CD38, making it another promising target in this disease.5

Blood cancer cells under the microscope: © stock_acc - stock.adobe.com

Blood cancer cells under the microscope: © stock_acc - stock.adobe.com

Potential New Targets for AML Treatment


CD4 is expressed at high levels on M4 and M5 AML subtypes, and its expression appears to correlate with blast percentage.10 As hematopoietic progenitor stem cells and nonhematopoietic cells do not express CD4, it offers promise as a target for certain patients.


An in vivo study showed that targeting CD93 had minimal damage to hematopoietic cells; however, there was notable on-target, off-tumor injury to endothelial cells where CD93 is expressed.5 A 2021 study identified that NOT gating strategies could ameliorate these endothelial toxicities, which would make CD93 a better target.11


Mesothelin, or MSLN, is not present during normal hematopoiesis but is highly expressed in AML cells, making it a potential target for CAR T-cell therapy that can protect against hematopoiesis. In vitro and xenograft models also showed that MSLN could be successfully targeted with antibody-drug conjugates.12


Velasquez said that GRP78 is present in every cell but limited to the endoplasmic reticulum, where it is a “key chaperone of the unfolded protein response.”

“Whenever there is cellular stress, for example, in the tumor cell, because they are more resistant and GRP78 gets translated to the cell surface. Then you get an antigen that, in theory, is present on the cell surface of your tumor but not on your normal cells,” Velasquez explained.

Velasquez and her team found that there was differential gene overexpression of GRP78 compared with normal CD34 cells and that regardless of whether the AML was a new diagnosis or relapse, the expression was present. This allowed the research team to create a CAR specific to GRP78.

“It is a second-generation CAR. The normal structure of a CAR is an antigen recognition domain that generally is derived from an antibody, and then you have several other regions, including a hinge, transmembrane, domain, costimulatory domain, and activation domain. In this case, we use a 13-mer peptide, a very short peptide to try to recognize GRP78,” Velasquez said.

“In the end, the 1 with a single peptide repeat worked better, so we tested it in vivo. When we tested it in vivo, we saw that there was tumor control, there was transient control, but it was not that great. Towards the end of the 50 days or so, the tumors would come back, and they were GRP78-positive and there were no T cells, so we found a way of using dasatinib [Sprycel], trying to retain the GRP78 intracellularly to try to dampen that problem and try to have the T cells persist a little bit longer. So that worked out,” Velasquez added.

Looking Ahead

The field of treatment in AML, especially cell therapies, has evolved quickly and continues to do so, and there is plenty of room for progress.

“Obviously, within the last, perhaps 5 to 6 years, AML therapy has remarkably changed. We used to use the ‘7 +3’ and then we had some HMA. But now we have a lot of combinations and quite a [number of] FDA-approved drugs. The field is obviously expanding,” said Musa Yilmaz, MD, faculty member in the leukemia department at MD Anderson Cancer center, in an interview with Targeted OncologyTM.

1. Atilla E, Benabdellah K. The black hole: CAR T cell therapy in AML. Cancers (Basel). 2023;15(10):2713. Published 2023 May 11. doi:10.3390/cancers15102713
2. CAR T-cell therapy. MD Anderson Cancer Center. Accessed April 17, 2024. https://tinyurl.com/43cuu5nv
3. Baulu E, Gardet C, Chuvin N, Depil S. TCR-engineered T cell therapy in solid tumors: State of the art and perspectives. Sci Adv. 2023;9(7):eadf3700. doi:10.1126/sciadv.adf3700
4. Zhang L, Meng Y, Yao H, et al. CAR-NK cells for acute myeloid leukemia immunotherapy: past, present and future. Am J Cancer Res. 2023;13(11):5559-5576. Published 2023 Nov 15.
5. Shao R, Li Z, Xin H, et al. Biomarkers as targets for CAR-T/NK cell therapy in AML. Biomark Res. 2023;11(1):65. Published 2023 Jun 17. doi:10.1186/s40364-023-00501-9
6. Gomes-Silva D, Atilla E, Atilla PA, et al. CD7 CAR T cells for the therapy of acute myeloid leukemia. Mol Ther. 2019;27(1):272-280. doi:10.1016/j.ymthe.2018.10.001
7. Sauer T, Parikh K, Sharma S, et al. CD70-specific CAR T cells have potent activity against acute myeloid leukemia without HSC toxicity. Blood. 2021;138(4):318-330. doi:10.1182/blood.2020008221
8. Clinical study of CLL1 CAR-T cells in the treatment of hematological malignancies. ClinicalTrials.gov. Updated February 23, 2022. Accessed April 16, 2024. https://clinicaltrials.gov/study/NCT05252572
9. Anti-CLL1 CAR T-cell therapy in CLL1 positive relapsed/refractory acute myeloid leukemia (AML). ClinicalTrials.gov. Updated May 13, 2021. Accessed April 16, 2024. https://clinicaltrials.gov/study/NCT04884984
10. Salman H, Pinz KG, Wada M, et al. Preclinical targeting of human acute myeloid leukemia using CD4-specific chimeric antigen receptor (CAR) T cells and NK cells. J Cancer. 2019;10(18):4408-4419. Published 2019 Jul 23. doi:10.7150/jca.28952
11. Richards R, Zhao F, Freitas K, et al. NOT-gated CD93 CAR T cells effectively target AML with minimized endothelial cross-reactivity. Blood Cancer Discov. 2021;2(6):648-665. Published 2021 Sep 16. doi:10.1158/2643-3230.BCD-20-0208
12. Kaeding A, Barwe S, Gopalakrishnapillai, A, et al. Mesothelin is a novel cell surface disease marker and potential therapeutic target in acute myeloid leukemia. Blood Adv. 2021;5(9):2350-2361. doi:10.1182/bloodadvances.2021004424
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