AML Patients in Minimal Residual Disease Vaccinated with a Novel Generation of Fast Dendritic Cells Expressing WT-1 and PRAME Mount Specific Immune Responses That Relate to Clinical Outcome

AML is frequently diagnosed in elderly patients, with a median age of 69.  Many older patients cannot tolerate intensive chemotherapy and/or stem cell transplantation, making curative treatment difficult and rates of early relapse high. Immunotherapy with dendritic cell (DC) vaccines after chemotherapy was shown by others to provide clinical benefit to some AML patients (van Tendeloo et al. 2010).

Here we report results in four AML patients receiving DC vaccines targeting the antigens Wilm’s tumor-1 (WT-1) and preferentially expressed antigen in melanoma (PRAME), applied in compassionate use, employing new generation monocyte-derived fast DCs, matured with a cocktail containing the TLR7/8 ligand R848. The mature DCs show high expression of CD83, strong up-regulation of HLA-DR and co-stimulatory molecules, down-regulation of CD14 and polarized release of IL-12p70, with no or low IL-10 secretion, upon T cell encounter.

After informed consent and hematopoietic recovery from chemotherapy, mononuclear cells were collected by apheresis and mature DC vaccines were prepared to separately express full length mRNA encoding the two target antigens (Subklewe et al. Cancer Immunol. Immunother. 2014). DCs were administered intradermally, once weekly for 4 wks, at wk6 and then on a monthly basis. Blood and bone marrow (BM) samples were collected throughout treatment. Minimal residual disease (MRD) was measured in BM and blood by quantitative PCR of WT-1 expression and BM was monitored by morphology.   

Table 1 summarizes the salient features of the patients, treatment parameters, MRD monitoring and initial immune response assessment. DTH reactions were detected in all patients challenged with DCs at wk6. Immune responses of CD4 and CD8 T cells demonstrating intracellular interferon gamma (IFNg) expression were assessed by flow cytometry of PBL stimulated overnight with peptides spanning WT-1, PRAME, and hTERT and survivin as vaccine-unrelated antigens. Responses were scored positive when two-fold or greater frequencies of IFNg-expressing T cells were found compared to unstimulated controls.

Patient (Pt.)CU030 and Pt.CU031 showed CD4 and CD8 responses to different test antigens. Pt.CU030 displayed strong and persistent CD8 responses to PRAME and a surprising increase in hTERT reactivity, potentially representing epitope spreading. The pt. continues to receive monthly vaccination and displays a low fluctuating WT-1 PCR signal in BM but no signal is seen in blood at wk61 after start of vaccination. Pt.CU031 displayed WT-1-specific immune responses until wk37 when responses decreased and WT-1 PCR signals increased in BM. The pt. developed Bell’s palsy and immune responses were no longer detected after cortisone therapy. WT-1 signals then increased strongly in BM, accompanied by an increase of blasts. Pt. CU033 had no significant T cell response during 9 months (m) of vaccination. WT-1 signals now increase slowly in BM but relapse cannot be confirmed by morphology and WT-1 PCR remains negative in blood. Pt.CU040 has only received DC vaccines for 5 m, remains in morphological remission and immune response and MRD monitoring are ongoing.

These results show that fast, TLR-polarized DCs induce or enhance specific T cell responses in elderly and undertreated AML patients, with individual strengths and specificities. Preliminary assessments suggest that changes in MRD are related to increase or loss of vaccine-associated immune responses.

Table 1: Characteristics of AML patients receiving DC vaccines

Patient	CU030	CU031	CU033	CU040
Age	57	50	68	73
Sex	f	m	f	f
AML Classification	M4	M2	M1	M1
Risk Classification	intermed	intermed	intermed	good
Chemotherapy cycles Induction/Consolidation	2/0	2/4	2/0	2/0
Time between chemo-therapy and vaccination	5 m	8 m	3 m	7 m
Months of vaccination as of (08/2015)	16 m	10 m	9 m	5 m
DTH responses at w6 toWT-1/PRAME DC challenge	pos/pos	pos/pos	pos/pos	pos/pos
IFNg-positive   T cell responses to overlapping peptides of WT-1, PRAME, hTERT, and Survivin	Strong  and persistent CD8 responses to PRAME and hTERT	Early CD4 & CD8 responses to WT-1; decrease at wk37; full loss after cortisone therapy	No significant responses detected up to wk33	To be done after acquisition of further samples
MRD (WT-1 PCR) in BM/blood	fluctuating low /neg	rapid increase after cortisone /pos	slow increase /neg	ongoing
BM morphology (most recent test)	neg	pos	neg	neg
Time since completion of chemotherapy	21 m	18 m	12 m	12 m