Induction Of In Vivo Oligoclonal T Cell Expansion and Specific In Vitro T Cell Responses Following K562/GM-CSF Vaccination Of MDS Patients
Authors: Prince Gabrielle , Thoburn Chris , Warlick Erica D. , Hess Allan D. , Smith B. Douglas 
Published: October 2019

Abstract

Background
Myelodysplatic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell malignancies, characterized by the ineffective hematopoiesis, and risk of progression to acute myeloid leukemia. Allogeneic stem cell transplant (SCT) remains the only potentially curative therapy, but toxicities limit its application in older adults. Vaccination strategies have been developed to modulate the immune system, ideally with less toxicity. We present results from a pilot study of single agent K562/GM-CSF whole cell vaccination in MDS patients (pts).

Methods
Poor-risk or transfusion-dependent MDS pts (n=5) were enrolled and received K562/GM-CSF whole cell vaccine (1 X 108 cells) every 3 weeks for 4 cycles followed by a booster vaccination 8 weeks later. Eligible pts could receive supportive care only for the 2 months preceding study entry, have no history of SCT, and not be on immunosuppressants. Blood, bone marrow, and skin biopsies were taken prior to vaccination, day 4 following first vaccine, and at 4 weeks after the booster vaccination. Diversity of the T cell compartment was assessed in the blood, marrow and skin using the TCRExpress Quantitative Analysis Kit (Biomed Immunotech, Tampa FL). Fragment length analysis was performed by the DNA Analysis Facility. In vitro stimulation studies for proliferation and cytokine production were performed using peripheral blood monocytes (PBMCs) for the pt with the best clinical response (>4 years). Proliferation of banked PBMCs, collected prior to every vaccination and stimulated with the vaccine, was assessed using 3H-Thymidine. The cytokines generated were measured using a multiplexed bead based immunoassay (Human 17-plex bioplex pro-panel) (Biorad, Hercules CA).

Results
All pts tolerated the vaccinations well with localized skin reactions being common. Clinically, 2 pts normalized their blood counts and became transfusion independent following the immunotherapy. One responding pt (CMML) remained stable without need for medical intervention for 4 yrs and only recently showed progression. T cell spectratyping indicated oligocolonal populations of T cells in post-vaccination skin, blood and marrow samples. T cells infiltrating the skin were tracked to the marrow. Interestingly, the best clinical responder demonstrated the most restricted skewed repertoire with a significant number of oligoclonal T cells tracking from skin to marrow (n=5). The marrow also had infiltrates of oligoclonal T cells not detected in the post-vaccination skin. Further, this skewed repertoire was absent when the pt relapsed. Proliferation assays measuring this pt’s T cell cytokine production in response to the vaccine cells in vitro, showed increased levels of IL-2, IL-13 and IL-5 that were suppressed or not produced by the time of the 4th vaccination. Inversely, IL-6, IL-10, IL-17, IL-1beta, MIP-1beta and TNF alpha levels increased throughout. The expected proliferative “boost” was seen with the initiation of the booster vaccine series at the time of progression, and co-culture of the pt’s lymphocytes with the vaccine cells suppressed the ability of the vaccine cells to produce GM-CSF in vitro. The ability to suppress GM-CSF production decreased during therapy and the pt’s lymphocytes had no effect on GM-CSF production by the vaccine at the end of the immunotherapy.

Conclusions
All pts showed T cell skewing by spectratyping analysis, suggesting that each had a change in T cell proliferation patterns in response to the vaccine. One pt had a significant clinical response and the most specific T cell response by spectratyping to the original vaccine, followed by the absence of these cells in the marrow at the time of progression. This suggests that an immune response may have stabilized his disease and progression was associated with loss of this T cell population. Proliferation studies suggest that the lymphocytes recognized the vaccine. Lastly, GM-CSF levels produced by the vaccine were decreased during the vaccination cycles suggesting that the pt’s lymphocytes and/or tumor had a suppressive effect on the vaccine cells. It is unclear if the GM-CSF suppression was essential, detrimental, or unrelated to the pt’s clinical response. Further study of the T cells patterns in these pts may elucidate details of the immune response that are integral to clinical responses.

Disclosures:
No relevant conflicts of interest to declare.