High-Dose RHAMM-R3 Peptide Peptide Vaccination for Patients with Vaccination for Patients with Acute Myeloid Leukemia (AML), Myelodysplastic Syndrome (MDS), Multiple Myeloma (MM) and Chronic Lymphocytic Leukemia (CLL)

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2911-2911 ◽  
Author(s):  
Jochen Greiner ◽  
Anita Schmitt ◽  
Krzysztof Giannopoulos ◽  
Isabel Funk ◽  
Marta Heyduk ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
T Cells ◽  
Regulatory T Cells ◽  
Immune Responses ◽  
Residual Disease ◽  
Lymphocytic Leukemia ◽  
High Dose ◽  
Clinical Effects ◽  
Peptide Vaccination

Abstract We have demonstrated immunological responses and positive clinical effects of a peptide vaccination for patients with AML, MDS, MM and CLL with a limited tumor load or a minimal residual disease over-expressing RHAMM using 300 μg RHAMM-R3 peptide (Schmitt et al., Blood 2008; Giannopoulos et al., abstract submitted). To date, 26 patients were enrolled in this clinical peptide vaccination trial. Here, we report on the second cohort of nine patients with AML, MDS and MM vaccinated with a higher peptide dose (1000 μg RHAMM-R3 peptide). The vaccine was given four times at a biweekly interval and GM-CSF was added for five days each vaccination. Similar to the patients vaccinated with 300 μg peptide only mild drug-related adverse events were observed such as erythema and induration of the skin. Immunomonitoring was performed using ELISpot assays for Interferon gamma and Granzyme B, tetramer-based flow cytometry and chromium release assays. Moreover, the frequency of regulatory T cells was quantified at different time points of vaccination. In this second cohort of patients treated with 1,000 μg peptide we detected specific immune responses in a lower frequency (4/9 patients) in contrast to patients in the 300 μg cohort (7/10 patients). In these patients with immune responses we found an increase of CD8+/HLA-A2/RHAMM-R3 tetramer+/CD45RA+/CCR7−/CD27−/CD28− effector T cells in flow cytometry in accordance with an increase of R3-specific CD8+ T cells in ELISpot assays. Two patients with positive immune responses showed a significant decrease of regulatory T cells. One patient without positive immune and clinical effects showed an increase of the frequency of regulatory T cells (5.03% to 15.9%). Three out of nine patients treated with 1,000 μg showed positive clinical effects: One patient with MDS RAEB-2 showed a reduction of leukemic blasts in the bone morrow to lower than 5%, one MDS patient achieved a normalization of the peripheral blood counts and one patient with multiple myeloma experienced a reduction of light chain in serum. The patients in the 300 μg cohort showed also a higher frequency of positive clinical effects (5 out of 10 patients). Taken together, RHAMM-R3 peptide vaccination induced both immunological and clinical responses. Therefore, RHAMM constitutes a promising structure for further targeted immunotherapies in patients with different hematological malignancies. However, higher doses of peptide do not improve the frequency and intensity of immune responses in this clinical trial.

Co-Existing Serological Immune Responses against RHAMM Might Be a Prerequisite for Strong Cellular Immune Responses of CD8-Positive T Cells in RHAMM-R3 Peptide Vaccination for Patients with Different Hematological Malignancies.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 3671-3671
Author(s):  
Jochen Greiner ◽  
Susanne Hofmann ◽  
Krzysztof Giannopoulos ◽  
Markus Rojewski ◽  
Anna Babiak ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
T Cell Responses ◽  
High Dose ◽  
Clinical Effects ◽  
Peptide Vaccination ◽  
Cell Responses ◽  
Tetramer Staining

Abstract Abstract 3671 Poster Board III-607 For effective elimination of malignant cells by specific T cells a co-activation of CD4- and CD8-positive T cells might be important. We performed two RHAMM-R3 peptide vaccination trials using 300μg and 1000μg for patients with AML, MDS and multiple myeloma overexpressing RHAMM. Similar mild toxicity of both cohorts was found, only mild drug-related adverse events were observed such as erythema and induration of the skin. In the 300μg cohort we detected in 7/10 (70 %) patients specific immune responses and also positive clinical effects in 5/10 (50 %) patients. In the high dose peptide vaccination trial (1000μg peptide) 4/9 (44 %) patients showed positive immune responses. These patients showed an increase of CD8+RHAMM-R3 tetramer+/CD45RA+/CCR7-/CD27-/CD28- effector T cells and an increase of R3-specific CD8+ T cells. In the higher peptide dose cohort three patients showed positive clinical effects. However, higher doses of peptide do not improve the frequency and intensity of immune responses in this clinical trial and might induce immune tolerance. In this work, we investigated the co-existence of serological immune responses against RHAMM detected by a RHAMM-specific ELISA of patients with AML, MDS and multiple myeloma treated in these two peptide vaccination trials. We correlated these results to specific T cell responses of CD8-positive T cells measured by ELISpot assays for interferon gamma and Granzyme B, tetramer staining and chromium release assays. Moreover, these results were compared to the frequency of regulatory T cells. 4/19 patients have a positive serological immune response in ELISA assay, all of these patients developed also strong specific CD8-positive T cell responses during peptide vaccination detected by ELISpot assays and tetramer staining. As expected, peptide vaccination did not result in the induction of humoral immune responses. In further ELISA assays we measured IL-2 and IL-10 levels in the sera of the patients before and three weeks after four vaccinations. While IL-10 levels remained at a rather low level over the time of vaccination, we detected an increase of IL-2 up to the five-fold of the initial levels in four of ten patients. Moreover, we performed a proteome array to detect cytokine and chemokine regulation in sera of patients vaccinated in these two trials during and after RHAMM-R3 peptide vaccination. 36 cytokines, chemokines and acute phase proteins were measured and both cohorts vaccinated with different peptide doses were compared. Taken together, RHAMM-R3 peptide vaccination induced both immunological and clinical responses. Co-existence of immune responses of CD4-positive T cells against the target RHAMM seems to be important for an induction of strong immune responses of CD8-positive T cells. Disclosures: No relevant conflicts of interest to declare.

The Absence of Vasoactive Intestinal Peptide Augments Allo-Reactivity and the Anti-Viral Response in Bone Marrow Transplantation.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3267-3267
Author(s):  
Lauren T. Southerland ◽  
Jian-Ming Li ◽  
Sohrab Hossain ◽  
Cynthia Giver ◽  
Wayne Harris ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
T Cell ◽  
Immune Responses ◽  
Cd8 T Cells ◽  
Cell Response ◽  
Low Dose ◽  
T Cell Response ◽  
High Dose ◽  
Mcmv Infection

Abstract Background: The severe morbidity and mortality associated with bone marrow transplantation (BMT) is caused by uninhibited immune responses to alloantigen and suppressed immune responses to pathogens. Vasoactive Intestinal Peptide (VIP) is an immunomodulatory neuropeptide produced by T-cells and nerve fibers in peripheral lymphoid organs that suppresses immune responses by induction of tolerogenic dendritic cells. In order to determine the immunoregulatory effects of VIP, we examined T-cell immune responses to allo- and viral-antigens in VIP knockout (KO) mice and mouse BMT recipients of hematopoietic cells from VIP KO donors. Methods: VIP KO mice and VIP WT littermates were infected with lethal or sub-lethal doses (5 × 104− 5 × 105 PFU) of murine cytomegalovirus (mCMV) and the T-cell response to viral antigen was measured by flow cytometry for mCMV peptide-MHC class 1-tetramer+ CD8+ T-cells. We transplanted 5 × 106 BM plus 1 × 106 splenocytes (SP) either from VIP KO or VIP WT donors in an C57BL/6 to F1(BL/6 × Balb/c) allo-BMT model and assessed survival, GvHD, donor T-cell expansion, chimerism, and response to mCMV vaccination and mCMV infection. Results: B-cell, αβ and γδ T-cell, CD8+ T-cell, CD11b+ myeloid cell, and dendritic cell numbers were equivalent between VIP KO and WT mice, while VIP KO mice had higher number of CD4+ and CD4+CD62L+CD25+ T-cells. Non-transplanted VIP KO mice survived mCMV infection better compared to VIP WT, with a brisker anti-viral T-cell response in the blood. In the allogeneic BMT setting, recipients of VIP KO BM plus VIP KO SP had more weight loss and lower (40%) 100 day post-transplant survival compared to the recipients of VIP KO BM plus WT SP (80% survival), recipients of WT BM plus KO SP (100% survival), and recipients of WT BM plus WT SP (80% survival). Recipients of VIP KO grafts had a significantly greater anti-mCMV response that peaked four days earlier than the tetramer response of mice transplanted with WT cells. This increased anti-viral response to vaccination correlated with a greater and more rapid T-cell response to secondary viral challenge. Conclusions: These experiments suggest that the absence of all VIP in the body, or the absence of VIP in a transplanted immune system, enhances anti-viral immunity and allo-immune responses. Modulation of the VIP pathway is a novel method to regulate post-transplant immunity. Figure 1: VIP knockout(KO) mice have an increased CMV tetramer response. VIP KO and VIP WT mice were infected (day 0) with either a sub-lethal low dose (5 × 10^4 PFU) or a lethal high dose (5 × 10^5 PFU) of CMV. Peripheral blood was stained for T cell markers and tetramer and analyzed by flow cytometry. On day 3, high dosed VIP KO mice had a higher number of tetramer positive CD8 T cells and better survival than WT mice (all high dose VIP WT died prior to day 10). VIP KO mice had a significant increase in tetramer positive CD8 T cells between days 3 and 10. *** p<0.01, difference between VIP KO and VIP WT littermate at designated dose level and day. Figure 1:. VIP knockout(KO) mice have an increased CMV tetramer response. VIP KO and VIP WT mice were infected (day 0) with either a sub-lethal low dose (5 × 10^4 PFU) or a lethal high dose (5 × 10^5 PFU) of CMV. Peripheral blood was stained for T cell markers and tetramer and analyzed by flow cytometry. On day 3, high dosed VIP KO mice had a higher number of tetramer positive CD8 T cells and better survival than WT mice (all high dose VIP WT died prior to day 10). VIP KO mice had a significant increase in tetramer positive CD8 T cells between days 3 and 10. *** p<0.01, difference between VIP KO and VIP WT littermate at designated dose level and day.

Final Results of Lower Dose Fludarabine (F) and Cyclophosphamide (C), and High Dose Rituximab (R), (FCR-Lite) for Patients with Untreated Chronic Lymphocytic Leukemia (CLL).

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2037-2037
Author(s):  
Ahmad A. Tarhini ◽  
S. Land ◽  
L. Pietragallo ◽  
A. Laman ◽  
M. Sulecki ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Response Rate ◽  
Residual Disease ◽  
Lymphocytic Leukemia ◽  
High Dose ◽  
Color Flow ◽  
Major Toxicity ◽  
Grade 3 ◽  
Age Range

Abstract Introduction Standard FCR therapy in untreated CLL patients (F-25 mg/m2 d1–3 q 4wk; C-250 mg/m2 d 1–3 q 4wk; R-500 mg/m2 d1 q 4wk for 6 cycles) was reported to have complete remissions (CR) of 70% and overall responses (OR) of 95% (J Clin Oncol2005;23:4079). The major toxicity was grade 3/4 neutropenia during 52% of treatment courses. One approach to decrease neutropenia without compromising efficacy could be by reducing the doses of F and C and increasing the dose of R. Methods We conducted a phase II study for previously untreated advanced CLL patients treated with FCR-Lite (F-20mg/m2 d1–3 q 4 wk; C-150 mg/m2 d1–3 q 4 wk; R-500mg/m2 d1 and d14 q 4wks; maintenance R-500 mg/m2 ×1 q 3 months until progression). A Simon two-stage design was used where 15 patients were accrued in the first stage and because of acceptable toxicity and response rate in stage I an additional 35 patients were treated. The primary endpoint was response rate. Results A total of 50 patients were entered into this study and 42 are currently evaluable. There were 29 male and 13 female patients with an age range of 36–85 years (median 58) treated with a total of 236 courses of FCR-Lite. All 42 patients were evaluable for toxicity. Grade 3/4 neutropenia occurred during 29 (12%) courses with two episodes of neutropenic fever. One patient had cellulitis, another had pneumonia (not neutropenic). Grade 3/4 thrombocytopenia occurred during 7 (3%) courses and grade III/IV anemia during 6 (2.5%) courses. Among the 40 evaluable patients for response, the CR rate was 85%, PR rate was 15% with an OR rate of 100%. All of the CR patients were tested by flow cytometry and had &lt;1% CD5+/CD19+ cells in their bone marrow after therapy. One patient with potential CR was excluded due to the absence of follow up bone marrow biopsy. Minimal residual disease (MRD) was tested by four color flow cytometry (sensitivity 0.01%) in 8 patients with CR (Genzyme Genetics Corp.). Seven had no evidence of MRD at 7, 8, 8, 14, 22, 25 and 30 months respectively, post CR, and one patient had 0.03% and 0.06% when tested at 12 and 18 months post CR respectively. Conclusions Our results in 42 patients suggest FCR-Lite is highly effective with considerably less grade 3/4 neutropenia than standard FCR. Complete responders had no detectable CD5+/CD19+ cells in their bone marrow following FCR-Lite. MRD testing is currently underway for all patients.

Monitoring Immune Responses Following Vaccination with Autologous Dendritic like Leukaemia Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1804-1804
Author(s):  
Matthias Klammer ◽  
Martin Waterfall ◽  
Kay Samuel ◽  
Charlotte Thomas ◽  
Peter R.E. Johnson ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Immune Responses ◽  
Minimal Residual Disease ◽  
Residual Disease ◽  
Fold Increase ◽  
Multi Centre Study ◽  
Successful Induction ◽  
Minimal Residual

Abstract It has been demonstrated that in a proportion of patients with Acute Myeloid Leukaemia (AML) their leukaemic blasts can be induced to differentiate into Dendritic Like Leukaemic Cells (DLLC) in vitro under the influence of GMCSF, IL-4 and TNF alpha. DLLC hold promise as cellular vaccines aimed at eradicating Minimal Residual Disease following successful induction of remission by chemotherapy. In A Phase I/II Multi-Centre Study we assessed leukaemic blasts of 21 patients with AML for their cytokine driven potential to differentiate into DLLC in vitro. 7 out of 21 (33%) patients showed DLLC conversion of their AML blasts based on morphological appearances and immunophenotype. 5 patients have completed chemotherapy and were then eligible to receive 4 escalating doses of subcutaneous DLLC vaccine. Immune responses to the administration of the vaccine are monitored by a combination of methods. The emergence of leukaemia specific T-cells following vaccination is demonstrated using Elispot assaying of Interferon gamma release in an in vitro re-stimulation assay. In WT1 expressing patients HLA-Tetramers allow to determine the frequency of WT1 specific T-cells. Regulatory T-cells (CD4/CD25 positive) are monitored by flow-cytometry. Monitoring of Minimal Residual Disease (MRD) is undertaken by means of real-time quantitative RT-PCR for leukaemia specific fusion transcripts or WT1 gene expression. 4 patients have so far completed the DLLC vaccination course. Vaccination was well tolerated with minimal side effects. A more than two-fold increase of leukaemia specific cytotoxicity was demonstrated following DLLC vaccination in one patient, while reduction of MRD was seen in several patients during the vaccination. The increase in CD4 /CD25 positive regulatory T-cells observed in several patients post vaccination may serve to dampen the induced anti-leukaemic immunity.

Peptide Vaccination Induces Dynamic Changes in CD4+ and CD8+ T Cell Subsets: Report on the First Peptide Vaccination Trial in Patients with Chronic Lymphocytic Leukemia (CLL)

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3159-3159 ◽  
Author(s):  
Krzysztof Giannopoulos ◽  
Malgorzata Kowal ◽  
Anna Dmoszynska ◽  
Jacek Rolinski ◽  
Kamila Mazurek ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Regulatory T Cells ◽  
Immune Responses ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
T Cell Subsets ◽  
Peptide Vaccination ◽  
Tetramer Staining ◽  
Induced Changes

Abstract There is an accumulation of in vivo (graft-versus-leukemia effect) and in vitro (spontaneous remissions after infections) data providing evidence that CLL might be effectively targeted by T-cell based immunotherapy. Earlier, we characterized the receptor for hyaluronic acid mediated motility (RHAMM) as antigen associated with proliferation and negative prognosis in CLL. We also demonstrated that RHAMM-derived epitope(R3)- primed T cells were able to lyse RHAMM+ target CLL cells. Therefore, we initiated a small phase I/II clinical trial with R3 peptide vaccination for patients with CLL. Six CLL patients in Binet stage 0 of the disease were vaccinated four times at a biweekly interval with HLA-A2 restricted RHAMM-derived epitope R3 (ILSLELMKL, 300μg/dose on day 3) emulsified in incomplete Freund’s adjuvant (IFA) with concomitant administration of GM-CSF (100μg/dose, days 1–5). R3-specific T-cell responses were assessed by tetramer staining and ELISPOT assays. T-cell subsets which play a role in regulation of immune responses including CD3+CD4+CD25hiCD127loFOXP3+ T regs, Th17, CD8+CD137+, CD8+CD103+ and IL-17 producing CD8+ T cells (CD8+IL-17+) were evaluated by flow cytometry. No severe adverse events greater than CTC Io skin toxicity could be observed. Four of six patients showed a reduction of WBC during vaccination. Although these WBC changes did not meet the NCI response criteria, we described these favorable hematological changes achieved in short period of immunotherapy as hematological improvement (defined as at least 20% reduction of WBC during vaccination). The immune responses were found in 5/6 patients as assessed by tetramer-staining (positive response defined as an increase of R3-specific CD8+ T cell frequency by more than 100% after vaccination) and confirmed in 4/5 as assessed by ELISPOT assay. Patients included in this study showed median Tregs frequency of 4.2%, range: 2.5–8%. There was no significant difference of Tregs percentages between patients who improved clinically when compared with non-responders (median 6.1% vs. 3.7%). Vaccination induced Tregs in 4 patients (2 non-responders and 2 responders). Two other patients who improved hematologically did not significantly change frequency of Tregs or even reduced it during vaccination (Figure 1). Median expression of CD103 on CD8+ T cells was 1.84%, range: 0.41–5.63%. In one non-responder, we observed an increase in frequency of CD103+CD8+ T-cells during vaccination from 1.46% to 2.56%. During vaccination, changes in CD8+CD103+ T cell subset did not correlate with the frequency of Tregs, nonetheless we could find an inverse correlation with inflammatory Th17 T cells (r2=−0.5, p&lt;0.05). We could find a correlation between the frequency of Tregs and activated CD8+CD69+ T cells (r2=0.51, p&lt;0.05). Interestingly, CD8+CD137+ cells correlated with CD8+IL-17+ T cells (r2=0.54, p&lt;0.05). In conclusion, peptide vaccination in CLL patients is safe and feasible to mount immune responses against the tumor antigen RHAMM. Most of patients benefited hematologically from vaccination. Although in some patients we observed an induction of tumor-specific T cells without induction of Tregs there is a rationale to add novel active agents against Tregs in future vaccination trials. Figure 1. Peptide vaccination induced changes in WBC, percentages of regulatory T cells (Tregs) as well as R3 specific tetramer ‘CD’ T cells (tetra) of A CLL patients. Patients B, C, E and F improved hematologically during vaccination. Figure 1. Peptide vaccination induced changes in WBC, percentages of regulatory T cells (Tregs) as well as R3 specific tetramer ‘CD’ T cells (tetra) of A CLL patients. Patients B, C, E and F improved hematologically during vaccination.

scholarly journals Imbalance between IL-17A-Producing Cells and Regulatory T Cells during Ischemic Stroke

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Yuehua Hu ◽  
Yanhua Zheng ◽  
Ya Wu ◽  
Bing Ni ◽  
Shugui Shi
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Ischemic Stroke ◽  
T Cell ◽  
Animal Models ◽  
Regulatory T Cells ◽  
Immune Responses ◽  
Real Time Pcr ◽  
Treg Cells ◽  
T Cell Subsets

Immune responses and inflammation are key elements in the pathogenesis of ischemic stroke (IS). Although the involvement of IL-17A in IS has been demonstrated using animal models, the involvement of IL-17A and IL-17-secreting T cell subsets in IS patients has not been verified, and whether the balance of Treg/IL-17-secreting T cells is altered in IS patients remains unknown. In the present study, we demonstrated that the proportion of peripheral Tregs and the levels of IL-10 and TGF-βwere reduced in patients with IS compared with controls using flow cytometry (FCM), real-time PCR, and ELISA assays. However, the proportions of Th17 andγδT cells, the primary IL-17A-secreting cells, increased dramatically, and these effects were accompanied by increases in the levels of IL-17A, IL-23, IL-6, and IL-1βin IS patients. These studies suggest that the increase in IL-17A-producing cells and decrease in Treg cells might contribute to the pathogenesis of IS. Manipulating the balance between Tregs and IL-17A-producing cells might be helpful for the treatment of IS.

Immunological and Clinical Responses in Patients with Acute Myeloid Leukemia (AML), Myelodysplastic Syndrome (MDS), Multiple Myeloma (MM) and Chronic Lymphocytic Leukemia (CLL) after RHAMM-R3 Peptide Vaccination.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1806-1806 ◽  
Author(s):  
Jochen Greiner ◽  
Anita Schmitt ◽  
Krzysztof Giannopoulos ◽  
Jinfei Chen ◽  
Marlies Goetz ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Immune Responses ◽  
Myeloid Leukemia ◽  
Hematological Malignancies ◽  
Lymphocytic Leukemia ◽  
Peptide Vaccination ◽  
Chromium Release ◽  
Clinical Responses ◽  
Acute Myeloid

Abstract We initiated a phase I/II R3 peptide vaccination for patients with acute myeloid leukemia (AML), myelodysplastic syndrome (MDS), multiple myeloma (MM) and chronic lymphocytic leukemia (CLL) overexpressing the receptor for hyaluronic acid mediated motility (RHAMM). RHAMM is a leukemia associated antigen (LAA) that is strongly expressed in several hematological malignancies and induces humoral and cellular immune responses. In this study, patients with AML, MDS, MM or CLL were included with RHAMM expression but with a limited tumor load or a minimal residual disease. To date, 25 patients were enrolled. The first 12 patients were vaccinated with 300 mcg and further patients with 1000 mcg R3 peptide emulsified with the incomplete Freund’s adjuvant. The vaccine was given four times at a biweekly interval and GM-CSF was added for five days each vaccination. Only mild drug-related adverse events were observed such as erythema and induration of the skin at the site of injection. Immunological analysis were performed using enzyme linked immunospot (ELISpot) assays for Interferon gamma and Granzyme B, tetramer staining and chromium release assays. We detected specific immune responses in 70% of patients. In most patients, we found an increase of CD8+/HLA-A2/RHAMM R3 tetramer+/CD45RA+/CCR7−/CD27−/CD28− effector T cells in flow cytometry in accordance with an increase of R3-specific CD8+ T cells in ELISpot assays. In chromium release assays, a specific lysis of RHAMM-positive leukemic blasts was shown. Moreover, we measured IL-2 and IL-10 levels in sera before and after vaccination. While IL-10 levels remained at a rather low level, we detected an increase of IL-2 in four of ten patients who showed also clinical responses. RHAMM transcripts in bone marrow and peripheral blood were quantified by real-time RT-PCR. Responding patients showed a decrease of RHAMM after vaccination. We detected positive clinical effects in several patients with myeloid disorders showing a reduction of blasts in the bone marrow. One MDS patient did not need any longer erythrocyte transfusions. Two patients with MM showed a reduction of free light chain serum levels. Taken together, RHAMM-R3 peptide vaccination induced both immunological and clinical responses. Therefore, RHAMM constitutes a promising structure for further targeted immunotherapies in patients with different hematological malignancies.

Regulatory T Cells Are Depleted in Low-Grade Lymphoma By the Combination of Local Low-Dose Radiation Followed By Intratumoral CpG-ODN

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1539-1539
Author(s):  
Debra K Czerwinski ◽  
Steven R Long ◽  
Michael Khodadoust ◽  
Matthew J. Frank ◽  
Michael P. Chu ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Regulatory T Cells ◽  
Immune Responses ◽  
Research Funding ◽  
Low Dose ◽  
Low Grade ◽  
Cpg Odn ◽  
Tfh Cells ◽  
Treated Site

Abstract BACKGROUND: Non-Hodgkin B cell lymphomas are often infiltrated by immune effector cells including T, NK and dendritic cells. But despite their presence within the tumor they fail to control tumor growth. Some T cells can even play a supportive role to maintain the tumor and prevent the function of anti-tumor immune cells. Regulatory T cells (TRegs) function normally to dampen immune responses and prevent auto-immunity by direct cell contact or by secreting suppressive cytokines i.e. TGF-b and Interleukin-10. Follicular Lymphoma tumor cells can induce the conversion of CD4 T cells into TRegs (1-3), thereby evading anti-tumor immune responses. Follicular helper T cells (Tfh), another class of regulatory T cells, are found within normal follicles of lymphoid organs. Migrating into the germinal center, they promote the survival and differentiation of follicular B cells. In lymphoma, Tfh cells can send a survival signal to tumor B cells through CD40L/CD40 interactions (4). STUDY: In an ongoing multi-center phase I clinical trial (NCT02266147), patients with previously untreated low-grade lymphoma receive low dose (2Gyx2) radiotherapy to a single, tumor site, followed by intratumoral injection into the same site of a CpG-ODN (SD-101, Dynavax Technologies), an immune-modulatory molecule that targets Toll-like Receptor 9 (TLR-9). Doses ranged from 1mg to 8mg per injection in successive cohorts. A fine needle aspirate (FNA) of the treated site is performed before, one week after the first injection, and 1 week after the 5th and final weekly injection of CpG-ODN. FNA samples are sent to a central site by overnight delivery in media with 5% fetal calf serum. They are processed within 24 hours and stained for flow cytometry with panels of antibodies to delineate T, B, NK, dendritic, myeloid cells, and their subsets. Antibodies against activation antigens, as well as T cell exhaustion, inhibition and function are also used to characterize these cells. To date, 12 patients have been entered into the dose escalation phase of this study: 10 follicular lymphoma, 1 chronic lymphocytic lymphoma and 1 small lymphocytic lymphoma. Nine of 12 patients had FNAs that yielded enough viable cells for evaluation pre and 1 week post treatment. RESULTS: Therapy has been well tolerated. Local injection site reactions and fever, the most common side effects, resolved by 48 hours. There have been no related SAEs. The treated site regressed in all patients per CT scan at 3 months post treatment. The early evaluation of untreated lesions documented 1 PR. The remaining patients have stable disease with all but one having systemic tumor regressions ranging from 14% to 28%. Seven out of 9 patients evaluable by flow cytometry had an increase in total T cells at the treated site 1 week following the first dose of CpG ranging from 18 to > 300% above baseline in both the CD4 and CD8 compartments. More interestingly, there was a reduction of TRegs as a percentage of total T cells in 7 of 9 patients (average reduction: 21.3 ± 10.6%). A single patient with an increase in untreated tumor burden of 15% showed no reduction in TRegs. There was also a reduction of Tfh cells in 8 of the 9 evaluable patients (average reduction: 87.2 ±7.2%). One patient with SLL had no Tfh cells pretreatment. Figure 1A and B shows a representative example of TReg and Tfh cell depletion within the same FL patient. CONCLUSION: Immune modulating therapies can be delivered directly into a tumor to minimize systemic toxicity and induce changes in the tumor microenvironment while potentially inducing a global anti-tumor immune response. In this case, the combination of intratumoral CpG and low-dose radiation reduced the proportion of TRegs and Tfh cells, thereby modulating their immune inhibitory effects and tumor growth promoting effects, respectively. This clinical trial is ongoing and open to accrual. Monitoring the cell populations in the tumor site by repetitive sampling and analysis by flow cytometry reveals the pharmocodynamic effects of anti-cancer therapies, especially those intended to trigger anti-tumor immune responses. This knowledge can be used to tailor therapies and to influence the choice and scheduling of combinations of agents designed to target specific cell populations. 1. Yang Z-Z, et al. Blood. 2007;110:537-44. 2. Ai WZ, et al. Int J Cancer. 2009 Jan 1;124(1):239-44. 3. Mittal S, et al. Blood. 2008;111:5359-70. 4. AmŽ-Thomas P, et al. Blood. 2015 Apr 9;125(15):2381-5 Figure 1. Figure 1. Disclosures Bartlett: Gilead: Consultancy, Research Funding; Janssen: Research Funding; Pharmacyclics: Research Funding; Genentech: Research Funding; Pfizer: Research Funding; Novartis: Research Funding; Millennium: Research Funding; Colgene: Research Funding; Medimmune: Research Funding; Kite: Research Funding; Insight: Research Funding; Seattle Genetics: Consultancy, Research Funding; MERC: Research Funding; Dynavax: Research Funding; Idera: Research Funding; Portola: Research Funding; Bristol Meyers Squibb: Research Funding; Infinity: Research Funding; LAM Theapeutics: Research Funding. Gordon:Northwestern University: Employment; Dr Leo I. Gordon: Patents & Royalties: Patent for gold nanoparticles pending. Coffman:Dynavax Technologies: Employment. Janssen:Dynavax Technologies: Employment. Levy:Bullet Biotechnology, Inc.: Consultancy.

scholarly journals Indirect induction of regulatory T cells accompanies immune responses during peptide vaccination of chronic lymphocytic leukaemia patients

2015 ◽  
Vol 174 (1) ◽  
pp. 155-157 ◽  
Author(s):  
Katarzyna Skorka ◽  
Joanna Zaleska ◽  
Malgorzata Zajac ◽  
Agnieszka Karczmarczyk ◽  
Waldemar Tomczak ◽  
...  
Keyword(s):  
T Cells ◽  
Regulatory T Cells ◽  
Immune Responses ◽  
Chronic Lymphocytic Leukaemia ◽  
Lymphocytic Leukaemia ◽  
Peptide Vaccination

Evaluation of MRD Status by Flow Cytometry and PCR in Multiple Myeloma Elderly Patients Receiving Autologous Stem Cell Transplantation After Bortezomib-Supplemented Mobilization and Conditioning

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4523-4523
Author(s):  
Immacolata Attolico ◽  
Roberta Nuccorini ◽  
Alberto Fragasso ◽  
Vincenzo Pavone ◽  
Pellegrino Musto ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Elderly Patients ◽  
Cell Transplantation ◽  
Autologous Stem Cell Transplantation ◽  
Residual Disease ◽  
Patient Specific ◽  
High Dose

Abstract Abstract 4523 Autologous Stem Cell Transplantation (ASCT) is still an option for eligible patients with Multiple Myeloma (MM). High-dose melphalan (HDM: 200mg/m∧2) is the recommended conditioning before ASCT, but synergistic effects of Bortezomib (BOR) and HDM have been reported in vitro and in vivo. PATIENTS AND METHODS: We evaluated in 56 MM fit elderly patients (median age 65 yrs), the feasibility and efficacy (also in terms of evaluation of minimal residual disease: MRD) of a strategy, combining BOR, Cyclophosfamide (CY) and dexamethasone (DEX) as induction and mobilizing therapy (CY-BOR), for ASCT, with conditioning including BOR-HD-MEL. The patients achieving at least PR after 4 CY-BOR courses, were mobilized with BOR and DEX standard schedule with CY 3g/m∧2 (day 8). The pts collecting at least 2.5×10∧6CD34+/kg underwent ASCT with HD-MEL (day-1) and BOR (1.0mg/m∧2 on -6,-3,+1,+4), followed by thalidomide consolidation until Relapse/Progression. The MRD has been prospectively evaluated both by using 4 colour flow cytometry (FC), and by using patient-specific probes, by ASO-PCR. The percentage of plasma cells (PCs) has been evaluated both in in PBSC harvested and in bone marrow, with CD38, CD45, CD56, CD138, CD19, CD27, CD28, CD117, kappa and lambda, along different steps of therapy. RESULTS: Of 44 pts evaluable for response before ASCT, 32 (73%) achieved 3PR and 30 (68%) were mobilized: 29 (66%) collected3 2.5×106CD34+/kg and 25 underwent ASCT. Median time for PMN engraftment was 11 days (range 10–13) and 14 (range12–20) for PLT>=20.000/mcl. We observed grade 3 neuropathy in 3 patients and pneumonia during induction in 2 patients. At day +180 from ASCT 23 are evaluable for response and 21 for MRD: 3 pts have progressive disease (PD), 2 pts have a PR, 4 pts have a VGPR, 10 pts a nCR and 4 pts a CR. Four colour FC, in order to detect clonal plasmacells (cPCs) along several steps of treatment, showed that 3 pts (14%) achieved MRD negativity: 1/21 pts achieved MRD negativity at day +180 (cPC <0.01%), being positive after induction and at day +90 after ASCT; two patients were MRD negative after induction (one developed positivity at day + 180 and relapsed at day +365 from ASCT; the other one became positive at day +90 after ASCT and, is in CR at 10 months from ASCT). In 5 patients we evaluated MRD by PCR with patient-specific probes. One patient achieved clearance of MRD after induction and still maintains negative of PCR at 27 months from ASCT: this patient had positivity of MRD by flow cytometry after induction and at 90 months from ASCT, then became negative and is in CR. One became PCR negative after ASCT: flow cytometry was negative too and the patient is in CR at 10 months from ASCT; the remaining three patients are PCR positive: two of them experienced progression of disease. CONCLUSIONS: ASCT with HDM and BOR is feasible in older patients, with very high RRs and without major toxicities. We need a longer follow up and a larger number of pts to assess if these results will translate in a benefit in terms of outcome. Disclosures: Fragasso: Mundipharma: Honoraria.

Sign in / Sign up

Export Citation Format

Share Document