Is three always greater than two?—azacitidine, venetoclax, and gilteritinib triple therapy in newly diagnosed and relapsed/refractory FLT3-mutant acute myeloid leukemia

Background

FLT3 is the most commonly mutated gene in adults with acute myeloid leukemia (AML) (1). The majority of FLT3 mutations are internal tandem duplications (ITD) with a smaller minority of point mutations in the tyrosine kinase domain (TKD), both resulting in constitutive activation of FLT3 signaling transduced through the MAPK and JAK/STAT pathways. In patients with FLT3-mutant AML, current standards of care include a FLT3 inhibitor in combination with intensive chemotherapy as frontline treatment: quizartinib for ITD mutations (2) and midostaurin for ITD or TKD mutations (3). In the relapsed/refractory (R/R) setting, the preferred treatment is monotherapy with the FLT3 inhibitor gilteritinib (4). Despite improvement in response rates from the addition of FLT3 inhibitors, resistance remains a ubiquitous clinical problem, and there is an unknown role for incorporating FLT3 targeted therapy into frontline treatment regimens for older patients who cannot tolerate intensive chemotherapy.

The combination of the DNA hypomethylating agent (HMA) azacitidine and the BCL-2 inhibitor venetoclax has emerged as the preferred therapy for older, frail, or medically unfit adults undergoing induction therapy for AML (5), or as a salvage regimen in patients lacking other intensive treatment options. BCL-2 is an anti-apoptotic protein whose inhibition has been shown to sensitize AML to cell death via apoptosis. There is a preclinical rationale for synergy between FLT3 inhibitors and venetoclax with reports that gilteritinib can increase BCL2 dependency and enhance venetoclax efficacy in vitro (6,7). Phase I/II data has shown the efficacy and tolerability of combination venetoclax and gilteritinib in R/R FLT3-mutant AML (8), but the role of triplet therapy with HMA plus venetoclax and gilteritinib remains unknown.

Trial design

In a recent issue of the Journal of Clinical Oncology, Short and colleagues published a single-center phase I/II investigation of azacitidine, venetoclax, and gilteritinib in both untreated and R/R FLT3-mutant AML (9). The phase I portion of the trial was conducted in R/R patients followed by phase II dose expansion cohorts with treatment naïve AML and R/R AML. Treatment was given with azacitidine 75 mg/m² intravenously or subcutaneously on days 1–7, venetoclax with ramp up to 400 mg daily orally on days 1–28 and gilteritinib orally on days 1–28. The dose of gilteritinib was 80 or 120 mg daily in the phase I cohort and 80 mg daily in the phase II cohorts.

Ten patients with R/R AML were enrolled in the phase I cohort to find the optimal dose of gilteritinib. Six were treated with gilteritinib 80 mg daily and four with gilteritinib 120 mg daily. There were no dose-limiting toxicities in the 80 mg arm while one patient in the 120 mg arm had grade 4 myelosuppression. Based on these data, 80 mg was chosen for the phase II dose expansion portion of the study.

Results in frontline cohort

The results of this trial are most notable for a complete remission (CR) rate of 90% (27/30 patients) in the frontline cohort, with a median age of 71 years (range, 18–86 years). Combined with patients who had complete remission with incomplete count recovery (CRi), there was a composite CR rate of 96%.

The outcomes of the frontline cohort reported by Short et al. compare favorably with those in the VIALE-A trial of azacitidine and venetoclax for treatment-naïve older adults with AML (albeit in a slightly older population with a median age of 76 years). There was a 96% composite CR rate in patients treated with the triplet vs. 66% in FLT3-mutant AML patients treated with azacitidine plus venetoclax (10). Responses were rapid, with 70% of patients showing morphologic response on day 14 marrow and 96% of patients achieving morphologic CR by the end of cycle 1, compared with a median time to first response of 1.3 months in VIALE-A. Importantly, the responses of patients treated with triplet therapy appear durable with 18-month overall survival (OS) of 72% compared to a median OS of 14.7 months in VIALE-A. Although more patients proceeded to hematopoietic stem cell transplant (HSCT) in the triplet trial than in VIALE-A, the 18-month OS rate for patients who did not receive HSCT was 79% vs. 63% for patients who received HSCT (P=0.22), suggesting that the OS benefit of the triplet over the doublet in the frontline does not depend on consolidative HSCT.

Notably, the results presented by Short et al. also compare favorably to results of intensive induction regimens. The 7+3 and midostaurin RATIFY trial only enrolled younger patients <60 and reported a CR rate of 58.9% (3). The similar AMLSG 16-10 trial enrolled patients up to 70 years old and reported a composite CR of 72% in patients 61–70 years old treated with 7+3 and midostaurin with 2-year OS of 57% (11). The 7+3 and quizartinib QuANTUM First enrolled patients up to 65 years old and reported a composite CR rate of 72% (2). At this time, the role of HMA plus venetoclax in induction therapy for younger or fit AML patients remains a topic of debate in the field. Nonetheless, the efficacy of triplet therapy suggested by this phase II data suggests that a randomized trial to compare survival outcomes for this patient population is indicated.

In recent years, the importance of measurable residual disease (MRD) in determining survival in AML has emerged, and the primary goal of modern frontline AML therapy has shifted to emphasize induction of remission with early eradication of MRD. By sensitive flow cytometry, 93% of patients ultimately achieved MRD negative status in this trial. However, it is known that MRD status by next-generation sequencing (NGS) detection of FLT3 ITD in FLT3-mutant AML predicts survival outcomes, and patients who are FLT3 ITD negative by NGS have lower risk of relapse and longer OS compared to patients who have even very low but detectable FLT3-ITD levels (12,13). In this study, 65% (13/20) of patients who were assessed for FLT3-ITD MRD by NGS achieved MRD negativity (FLT3-ITD frequency less than 5×10⁻⁵) by the end of cycle 4, suggesting deep molecular responses.

Results in the R/R cohort

In the R/R cohort (22 patients), outcomes were significantly worse than in the frontline cohort, likely due to adverse disease biology and prior treatment exposure. The median number of previous lines of therapy was 2, with 36% of patients having received a previous FLT3 inhibitor, 45% having received previous HMA plus venetoclax, and 27% having received previous HSCT. Reponses underperformed expectations, with modified composite CR rate [defined as combined CR/CRi/morphological leukemia-free state (MLFS) rate] of 68%, median relapse-free survival (RFS) 4.3 months, and median OS 5.8 months with a median follow up time of 30.7 months. The outcomes in the R/R cohort did not surpass previous results with gilteritinib monotherapy in patients at first relapse (modified composite CR 54%, median OS 9.3 months) (4). In a similarly pre-treated R/R population (median 2 lines of prior therapy), gilteritinib in combination with venetoclax yielded a modified composite CR rate of 75% and median OS 10.0 months (8). Investigation in a population at first relapse may reveal a greater efficacy of the triplet in the salvage setting. Indeed, a subgroup analysis showed a trend toward superior OS in patients who had not received prior treatment with HMA and venetoclax and/or gilteritinib compared to those who had (median OS 10.5 vs. 4.2 months, P=0.095), but it is currently unclear whether the triplet is superior to either HMA plus venetoclax, gilteritinib/venetoclax or even gilteritinib monotherapy.

Toxicity

There is understandable concern that triplet therapy with HMA, venetoclax, and a FLT3 inhibitor might have limited usefulness due to excessive myelosuppression (14). In this study, dose modification measures were taken to avoid excess toxicity. Bone marrow biopsy was performed on day 14 of cycle 1, and venetoclax was held if aplastic or if blasts <5%. For the frontline cohort only, gilteritinib was also held in this setting. For cycle 2 onward, azacitidine was given for 5 days, venetoclax for 7 days, and gilteritinib for 28 days. In the frontline cohort, 50% of patients had grade 3 infection and 33% had grade 3 febrile neutropenia. Only two patients in the frontline cohort had grade 4 or higher adverse events. There was significantly more toxicity in the R/R cohort, with 5 patients suffering grade 5 adverse events, all in the setting of progressive leukemia. The toxicity profile of the triplet regimen compares favorably to that of the azacitidine and venetoclax doublet published in VIALE-A, in which 64% of patients suffered grade 3 infections and 30% had grade 3 febrile neutropenia (5). This comparison highlights the importance of dose-modification to minimize toxicity while preserving efficacy.

Future directions

Overall, the results of this phase I/II trial of azacitidine, venetoclax and gilteritinib are encouraging and suggest a role for this triplet regimen in the management of adults with FLT3-mutant AML. However, there are several important questions that arise from the data.

What is the optimal dose of gilteritinib in the frontline setting?

The dose-finding phase I portion of this trial was only conducted in R/R patients, with the lowest effective dose of 80 mg being used for the subsequent frontline cohort. It is possible that treatment naïve patients might tolerate a higher dose of gilteritinib than R/R patients. The authors describe an ongoing dose-finding trial in the frontline population to determine optimal gilteritinib dosing in the frontline setting (NCT05520567).

Is the triplet superior to HMA plus venetoclax for older patients?

In the frontline cohort, the triplet regimen has very encouraging results with high response rates and durable remissions. Since the publication of the phase III VIALE-A trial, the doublet of HMA and venetoclax has been standard of care for older or medically frail adults with AML, including those with FLT3 mutations. Indeed, subsequent analysis of patients enrolled in VIALE-A showed no significant difference in outcomes between patients with mutated vs. wild-type FLT3 (10). In our institution and others, standard practice for these patients has been to treat upfront with a mutation-agnostic HMA plus venetoclax regimen and save targeted therapy with gilteritinib for the eventuality of relapse. The results described by Short et al. strongly suggest that addition of gilteritinib to HMA plus venetoclax might be superior treatment for FLT3-mutant patients.

Is the triplet superior to intensive chemotherapy plus FLT3 inhibitor for younger patients?

Based on the results of Short et al., younger patients who are currently treated with intensive chemotherapy plus quizartinib or midostaurin may have lower toxicity and better outcomes with an HMA, venetoclax, gilteritinib triplet. Randomized comparison of the triplet with standard of care regimens are needed to rigorously address this question.

What mechanisms mediate resistance to this triplet regimen?

The durability of response to triplet therapy in frontline patients with FLT3-mutant AML, with median RFS not been reached, suggests triplet therapy may effectively suppress some mechanisms of relapse. Further follow up will be important to understand patterns of relapse and mechanisms of resistance in patients treated with this triplet regimen. Among the frontline cohort there were 5 relapses, two of which were FLT3 positive. Based on work by our group and others on resistance to gilteritinib, we would expect mutations in the RAS/Mitogen-associated protein kinase pathway outside of FLT3 to be an important mechanism of resistance (15). Recent data from the BEAT-AML trial and others has led to the hypothesis that acquired monocytic differentiation may also be an important mechanism of resistance to venetoclax (16-18). We have performed single cell genomic analysis of patients treated with the combination gilteritinib and venetoclax, which shows upregulation of features of monocytic differentiation at the time of relapse (19). Longitudinal genetic and immunophenotypic profiling of patients treated with the triplet regimen will be necessary to understand the contributions of genetic evolution and cell state plasticity to treatment resistance.

In summary, the study of Short et al. is a single-center phase I/II trial investigating the safety and efficacy of the combination of azacitidine, venetoclax, and gilteritinib triplet therapy in adults with newly diagnosed or R/R FLT3-mutant AML. The frontline cohort showed very high rates of complete and durable remissions with acceptable toxicity, which suggests that this regimen should be investigated in randomized trials to determine its suitability as standard of care induction therapy in FLT3-mutant AML. The results in the small, heavily pre-treated R/R cohort do not compare favorably to standard therapies for R/R FLT3-mutant AML, but the efficacy of this regimen as a salvage option at first relapse remains unknown. As larger trials of this regimen progress, it will be important to identify mechanisms of treatment resistance, which are likely mediated by both genetic mutations and cell state plasticity.

Acknowledgments

Funding: None.

Footnote

Provenance and Peer Review: This article was commissioned by the editorial office, AME Clinical Trials Review. The article has undergone external peer review.

Peer Review File: Available at https://actr.amegroups.com/article/view/10.21037/actr-24-129/prf

Conflicts of Interest: Both authors have completed the ICMJE uniform disclosure form (available at https://actr.amegroups.com/article/view/10.21037/actr-24-129/coif). C.C.S. has received research support from Abbvie, served on an advisory board for Abbvie and provided educational talks for Astellas. The other author has no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

References

1. Papaemmanuil E, Gerstung M, Bullinger L, et al. Genomic Classification and Prognosis in Acute Myeloid Leukemia. N Engl J Med 2016;374:2209-21. [Crossref] [PubMed]
2. Erba HP, Montesinos P, Kim HJ, et al. Quizartinib plus chemotherapy in newly diagnosed patients with FLT3-internal-tandem-duplication-positive acute myeloid leukaemia (QuANTUM-First): a randomised, double-blind, placebo-controlled, phase 3 trial. Lancet 2023;401:1571-83. Erratum in: Lancet 2023;402:1328. [Crossref] [PubMed]
3. Stone RM, Mandrekar SJ, Sanford BL, et al. Midostaurin plus Chemotherapy for Acute Myeloid Leukemia with a FLT3 Mutation. N Engl J Med 2017;377:454-64. [Crossref] [PubMed]
4. Perl AE, Martinelli G, Cortes JE, et al. Gilteritinib or Chemotherapy for Relapsed or Refractory FLT3-Mutated AML. N Engl J Med 2019;381:1728-40. Erratum in: N Engl J Med 2022;386:1868. [Crossref] [PubMed]
5. DiNardo CD, Jonas BA, Pullarkat V, et al. Azacitidine and Venetoclax in Previously Untreated Acute Myeloid Leukemia. N Engl J Med 2020;383:617-29. [Crossref] [PubMed]
6. Zhu R, Li L, Nguyen B, et al. FLT3 tyrosine kinase inhibitors synergize with BCL-2 inhibition to eliminate FLT3/ITD acute leukemia cells through BIM activation. Signal Transduct Target Ther 2021;6:186. [Crossref] [PubMed]
7. Singh Mali R, Zhang Q, DeFilippis RA, et al. Venetoclax combines synergistically with FLT3 inhibition to effectively target leukemic cells in FLT3-ITD+ acute myeloid leukemia models. Haematologica 2021;106:1034-46. [Crossref] [PubMed]
8. Daver N, Perl AE, Maly J, et al. Venetoclax Plus Gilteritinib for FLT3-Mutated Relapsed/Refractory Acute Myeloid Leukemia. J Clin Oncol 2022;40:4048-59. [Crossref] [PubMed]
9. Short NJ, Daver N, Dinardo CD, et al. Azacitidine, Venetoclax, and Gilteritinib in Newly Diagnosed and Relapsed or Refractory FLT3-Mutated AML. J Clin Oncol 2024;42:1499-508. [Crossref] [PubMed]
10. Konopleva M, Thirman MJ, Pratz KW, et al. Impact of FLT3 Mutation on Outcomes after Venetoclax and Azacitidine for Patients with Treatment-Naïve Acute Myeloid Leukemia. Clin Cancer Res 2022;28:2744-52. [Crossref] [PubMed]
11. Döhner H, Weber D, Krzykalla J, et al. Midostaurin plus intensive chemotherapy for younger and older patients with AML and FLT3 internal tandem duplications. Blood Adv 2022;6:5345-55. [Crossref] [PubMed]
12. Grob T, Sanders MA, Vonk CM, et al. Prognostic Value of FLT3-Internal Tandem Duplication Residual Disease in Acute Myeloid Leukemia. J Clin Oncol 2023;41:756-65. [Crossref] [PubMed]
13. Dillon LW, Gui G, Ravindra N, et al. Measurable Residual FLT3 Internal Tandem Duplication Before Allogeneic Transplant for Acute Myeloid Leukemia. JAMA Oncol 2024;10:1104-10. [Crossref] [PubMed]
14. Yilmaz M, Kantarjian H, Short NJ, et al. Hypomethylating agent and venetoclax with FLT3 inhibitor “triplet” therapy in older/unfit patients with FLT3 mutated AML. Blood Cancer J 2022;12:77. [Crossref] [PubMed]
15. McMahon CM, Ferng T, Canaani J, et al. Clonal Selection with RAS Pathway Activation Mediates Secondary Clinical Resistance to Selective FLT3 Inhibition in Acute Myeloid Leukemia. Cancer Discov 2019;9:1050-63. [Crossref] [PubMed]
16. Bottomly D, Long N, Schultz AR, et al. Integrative analysis of drug response and clinical outcome in acute myeloid leukemia. Cancer Cell 2022;40:850-864.e9. [Crossref] [PubMed]
17. Kuusanmäki H, Leppä AM, Pölönen P, et al. Phenotype-based drug screening reveals association between venetoclax response and differentiation stage in acute myeloid leukemia. Haematologica 2020;105:708-20. [Crossref] [PubMed]
18. Pei S, Pollyea DA, Gustafson A, et al. Monocytic Subclones Confer Resistance to Venetoclax-Based Therapy in Patients with Acute Myeloid Leukemia. Cancer Discov 2020;10:536-51. [Crossref] [PubMed]
19. Kennedy VE, Peretz C, Lee P, et al. Multi-Omic Single-Cell Sequencing Reveals Genetic and Immunophenotypic Clonal Selection in Patients With FLT3-Mutated AML Treated With Gilteritinib/Venetoclax. Blood 2022;140:2244-6. [Crossref]