scholarly journals Association between BCL2, BCL2 E17, MCL1 and BAX Protein Expression, Bone Marrow Microenvironment Histological Features, Clinical Presentation, Therapeutic Outcome, and the Overall Survival in Newly Diagnosed Multiple Myeloma

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4716-4716
Author(s):  
Anita Skrtic ◽  
Ozren Jaksic ◽  
Slobodanka Ostojic ◽  
Mariastefania Antica ◽  
Petra Crnkovic ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Partial Response ◽  
Protein Expression ◽  
Cd8 T Cells ◽  
High Intensity ◽  
Overall Response Rate ◽  
Response Rate ◽  
Newly Diagnosed ◽  
Median Percentage

Abstract We evaluated the correlation between immunohistochemical BCL2, BCL2 E17, MCL1 (Bcl2-related protein family) and BAX (effector proapoptotic protein) expression and intensity of staining in plasma cells (PC) in MM patients (pts) who were eligible for autologous stem cell transplantation (ASCT) with DD4, CD8, and FOXP3 positive Tregs lymphocytes in bone marrow (BM) microenvironment, morphologic features of PC, clinical parameters, and outcomes. Immunohistochemical staining of BCL2, BCL2 E17, MCL1, BAX, CD4, CD8, FOXP3 and Ki67 was performed in BM biopsy of 36 newly diagnosed MM patients who were eligible for ASCT between 2012-2017 at University Hospital Merkur. Pearson's and Spearmans's coefficients of correlation, t-tests, one-way independent sample ANOVAs, mixed ANOVAs, moderation analyses, chi-square tests of independence and Cox's proportional hazards were used to analyze the data. Median age of pts was 57 years (95% CI 52-60), 15 pts were female with equal distribution in rISS I-III (12 pts). Twelve (33%) pts had high risk cytogenetics: del 13q34 (8 pts; 22%), del 17p (5 pts, 13,8%), t(4; 14) (4 pts; 11.1%); t(16;14) (4 pts; 11.1%); 4 pts (11.1%) had t(11;14). The median percentage of PC (PC ptc) in BM was 60 (95% CI 45 - 80); 25 pts (69.4%) had diffuse type of infiltration (TI) in BM; and 23 pts (63.8%) had high grade of PC differentiation (grade 2/3). The median percentage of Ki67 in PC was 3% of PC (95% CI 2 - 4, range 1 - 80). The median percentage of BCL2 positive PC in BM was 50 (95% CI 25 - 60) with high intensity of staining in 23 pts (63.8 %); BCL2 E17 positive PC in BM was 60 (95% CI 60 - 70) with high intensity of staining in 33 pts (91.6%); MCL1 positive PC in BM was 40 (95% CI 20 - 60) with high intensity of staining in 21 pts (58.3%); and BAX positive PC in BM was 17.5 (95% CI 6 - 40) with high intensity of staining in 18 pts (50%). Median CD4 positive T-cells was 8.5 (95% CI 4 - 18); CD8 T-cells 24 (95% CI 18 to 37); and FOXP3 Tregs 1 (95% CI 1 - 2, range 1 - 31). Nineteen pts (52.7 %) underwent tandem ASCT, 34 pts (94.4%) received VCD induction; 2 pts received MP induction; and 1 pts received VD induction. Overall response rate (≥partial response) to induction therapy was 94.4%; ≥ very good partial response rate was 80.5%. Overall response rate (≥partial response) to ASCT was 86.1%; complete response rate was 50%. Median overall survival time was 35.40 months (95% CI 26.61-165.39). Expression of BCL2, BCL2 E17, MCL1 and BAX proteins was significant and positively, moderately to strongly associated (p < 0.05), except for BCL2E17 and BAX association (r (36) = 0.319, p = 0.058). BCL2 and MCL-1 expression was more correlated when PC were well differentiated, grade 1 (p = .016). Higher PC pct was correlated with higher intensity of BLC2 staining (t (34) = 2.51, p = 0.017). Patients with Ki67 >10% had a higher PC pct in BM (t (34) = 2.04, p = 0.049). Higher grade of PC in BM correlated with higher PC pct in BM (t (34) = 2.63, p = 0.012). A higher MCL1 expression was found in MM with high-risk cytogenetic changes (t (34) = 2.09, p = 0.045). Cytoplasmic granular MCL1 staining had greater MLC 1 staining intensity than the diffuse TI (χ 2 (1) = 6.60, p = 0.017). Low BCL2, BCL2E17, MCL1 and BAX expression was found in MM stage rISS1 compared to rISS3 (p < 0.05), and low BCL2 and MCL-1 expression in MM stage rISS1 compared to ISS2 (p < 0.05). Patients with non-diffuse TI had worse response to ASCT compared to their response to induction therapy than patients with diffuse TI (χ 2 (2) = 6.39, p = 0.041). Although we observed high number of CD4 and CD8 T-cell in BM microenvironment and aberrant CD4:CD8 ratio in favor to CD8 positive T-cell, it had no impact on other variables in study. We found significant correlation between BCL2E17 expression and response to ASCT (r s = -0.417, p = 0.011), with higher protein expression being associated with worse outcomes. BCL2 contributed to worse survivability (χ 2 (1) = 7.18, p = 0.007), while BCL2 E17, MCL1 and BAX expression, as well as the number of CD4 and CD8 T-cells, did not predict survival (p > 0.05). Results showed aberrant expression of the BCL2 family members that may plays a role in application strategies for MM therapy. Bcl-2 inhibits Bax but we found aberrant BAX expression in MM that may be result of pre-condition or hormesis like responses in stressful micro-environments that serve to increase apoptotic resistance. Further investigation of the role Bcl2- proteins family in the biology of MM and its impact on clinical and therapeutic outcomes is warranted. Disclosures Jaksic: Roche, Oktal-Pharma/Celtrion, Sandoz: Consultancy, Honoraria.

scholarly journals Characterising the Immunological Microenvironment in Newly Diagnosed Multiple Myeloma Bone Marrow By Time of Flight Cytometry Reveals Abnormalities in Antigen Presenting and Effector Lymphocyte Populations with Prognostic Significance

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 58-58 ◽  
Author(s):  
Frances Seymour ◽  
Jamie Cavenagh ◽  
John G. Gribben
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Cd8 T Cells ◽  
Research Funding ◽  
Cytokine Production ◽  
Immune Surveillance ◽  
Cell Populations ◽  
Population Abundance ◽  
Newly Diagnosed ◽  
Time Point

Abstract Background Loss of immune surveillance is thought to contribute to disease progression and treatment resistance in a range of malignancies including multiple myeloma (MM). Understanding the degree and pattern of immunological abnormality present within the bone marrow microenvironment at the time of MM diagnosis is vital if we are to utilize emerging immunological therapeutic strategies successfully in MM. While immune checkpoint inhibition in the relapsed refractory setting has been disappointing, early intervention before immune subversion mechanisms have become established may enable more effective restoration of immunological disease control. Method Bone marrow samples from 18 patients with newly diagnosed, untreated, myeloma (NDMM) and 9 age matched bone marrow controls were labelled with metal-antibody conjugates and assessed by time of flight cytometry using the CyTOF platform. Expression of 36 protein targets including markers of proliferation, degranulation and cytokine production alongside phenotyping and viability markers were measured at the single cell level. Differences in the abundance and function of distinct cellular populations were assessed using the unsupervised, automated CITRUS algorithm with the goal to identify novel cell populations. Results We observed a decrease in three key populations in NDMM; CD4 T cells with an effector phenotype (CD4EF), CD8 T cells with an IL2 producing effector phenotype (CD8EF), and dendritic cells. The expected elevation in malignant plasma cells was also seen and characterized. Interestingly CD8 T cells with a cytotoxic phenotype were not decreased. The dendritic cell (DC) population contained two distinct subpopulations characterized by their level of CD16 expression. The CD16 positive population (DC16) expressed a range of activating receptors and cytokines while the CD16 negative population (DCTOL) exhibited strong Ki67 expression. Within NDMM samples the DC16 population had stronger expression of PDL1 (p=0.0349) and loss of TIM3 (p=0.0122) and 2B4 (p=0.0148) compared to controls suggesting that this subset is less functionally active in NDMM. The DCTOL population had a similar increase in PDL1 (p=0.0065) and loss of TIM3 (p=0.0165) but also had a shift towards CD107a (p=0.0014) and perforin (p=0.0196) expression, suggesting a tolerogenic role for this subset in myeloma. Within the CD4EF subset NDMM samples exhibited reduction in Ki67 (p=0.0054) compared to controls, suggesting that the decrease in population abundance might be due to loss of proliferation. Furthermore, the NDMM population had increased expression of TGFβ (p=0.0027) and FoxP3 (p=0.0409) suggesting that those cells that are present may have a regulatory role. The CD8EF population also showed reduction of Ki67 (p=0.0494) in NDMM compared to control samples. This was accompanied by a loss of DNAM1 (p=0.0096), suggesting a loss of co-stimulatory capacity, alongside elevations in TGFb (p=0.0022) suggesting a pro-tumor cytokine shift. A broad spread of cellular abundance levels was noted in NDMM compared to controls which led us to investigate whether differences in cell population abundance was associated with survival. This was seen for the CD8EF population, with higher abundance correlating with longer survival (r=0.6643, p=0.0026). Individuals with higher abundance of both the CD8EF population and the DC16 population had a reduction in relapse and death in the first 36 months following diagnosis (p=0.0366). Conclusions This data demonstrates that even at the early time point of myeloma diagnosis there is evidence of both numerical and functional defects in key cell populations involved in antigen presentation and anti-tumor activity. We propose that ineffective antigen presentation by PDL1 expressing DC populations results in poorly proliferative CD8 and CD4 effector populations with pro-tumor cytokine production. The high FoxP3 expressing CD4 population may also have a regulatory role. This data highlights PDL1 as an important therapeutic target in NDMM where it may have a role in restoring immune surveillance at an early disease time point. Harnessing emerging deep profiling technology to identify patterns of immunological change across multiple cellular subsets within one individual may enable us to identify immune signatures which predict outcome and response to treatment. Disclosures Seymour: Celgene: Research Funding. Cavenagh:Celgene: Honoraria, Research Funding, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Takeda: Research Funding, Speakers Bureau; Janssen: Honoraria, Speakers Bureau; Amgen: Honoraria, Speakers Bureau. Gribben:Kite: Honoraria; Roche: Honoraria; TG Therapeutics: Honoraria; Pharmacyclics: Honoraria; Cancer Research UK: Research Funding; Medical Research Council: Research Funding; Unum: Equity Ownership; Wellcome Trust: Research Funding; Acerta Pharma: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; NIH: Research Funding; Novartis: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Abbvie: Honoraria.

Patients with Newly Diagnosed Multiple Myeloma and a Gain of the Long Arms of Chromosome 1 Have Inferior Outcomes, Including Early Onset Extramedullary Disease, Despite the Use of Novel Triplet Regimens

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 4996-4996
Author(s):  
Noa Biran ◽  
Vesna Najfeld ◽  
Emily Bagiella ◽  
Sundar Jagannath ◽  
Ajai Chari
Keyword(s):  
Spinal Cord ◽  
Multiple Myeloma ◽  
Bone Marrow ◽  
High Risk ◽  
Overall Response Rate ◽  
Response Rate ◽  
Chromosome 1 ◽  
Newly Diagnosed ◽  
Inclusion Criteria ◽  
Novel Agent

Abstract Abstract 4996 BACKGROUND: In patients with newly diagnosed multiple myeloma (MM), a gain of the long arms of chromosome 1 is found in approximately 20% of patients by cytogenetics and 40% by FISH. A few studies have not found these abnormalities to be predictive of inferior progression free survival (PFS) and overall survival (OS) on multivariate analysis. The many studies that have were prior to the use of novel therapies in induction. Currently, testing for a gain or amplification of 1q21 locus is not included in the high risk genetic abnormalities by the International Myeloma Working Group (IMWG). Recently, two groups have reported inferior outcomes with bortezomib based induction regimens followed by high dose melphalan consolidation and maintenance therapy. The gain of 1q21 was either detected by cytogenetics or gene expression profiling (GEP70) at the University of Arkansas (Waheed et al, Cancer 2011) or by FISH in CD138 selected cells by the HOVON group (Neben et al, JCO 2012). However, the outcomes of these patients when treated with a bortezomib, lenalidomide, and dexamethasone induction regimen, which is associated with an overall response rate (ORR) of 100% and VGPR or better rate of 74% is unknown (Richardson et al. Blood 2010). Here, we describe the poor outcomes of patients with newly diagnosed MM with gain of 1q21 by FISH in unselected bone marrow aspirates despite novel agent triplet therapy. METHODS: The inclusion criteria for this IRB approved retrospective study were patients with symptomatic MM starting in June of 2008 who had a gain of 1q21 by FISH in 200 bone marrow interphase cells at the time of diagnosis. Patients with 1q amplification had at least 3 or more copies. PFS and OS were calculated by Kaplan-Meier analyses. RESULTS: 23 patients met the inclusion criteria. The median age was 59. 7 (range 46–71) and twenty patients (87%) were DS III at diagnosis, 13/23(57%) were ISS Stage 3. 6/23 (26%) had hypercalcemia, 8/23 (35%) had renal insufficiency, and 19/23 (83%) had anemia. Lytic lesions were present in 17/23 (74%) of patients at diagnosis. Of note, while 4 patients had deletion 13 by cytogenetics only 2 patients had other high risk findings, one with t(4;14) and another with deletion 17. All patients were treated with novel agent induction therapy. 19/23 (83%) were treated with triplet regimens (bortezomib, dexamethasone, and either lenalidomide or cyclophosphamide i. e. VRD or VCD). Disappointingly, primary induction failure, defined by PD or SD after 3–4 cycles, was observed in 30% of all patients. Of the 17 patients who received upfront triplet VRD or VCD therapy (with or without HDM plus SCR) the overall response rate (ORR) was only 77% (13/17), and 47% (8/17) achieved VGPR or better. More specifically, 3/17 (18%) had CR, 5/17 (29%) had VGPR, 5/17 (29%) had PR, 1/17 (6%) had SD and 1/19 (6%) had PD. Of the 4 patients who received novel doublet therapy (VD or RD) upfront, only 1 had a VGPR, one had SD and 2 had PD. The responses noted were not very durable, with a median PFS of 14 months. Although 3 patients have died (after 11–13 months from diagnosis), the median OS has not been reached with a median follow up of 13 months. Plasmacytomas of the bone with soft tissue expansion were present in 48% of the patients and 3 of 23 (13%) had spinal cord compression. Extra-osseous MM within 3 months of diagnosis was observed in 5/23 (22%) patients and 4/5 did not have any other high risk cytogenetics. Three patients (13%) had CNS MM at median of 5. 3 months after diagnosis. One patient had concomitant parenchymal brain lesions, myleomatous meningitis, and intra spinal cord disease. CONCLUSIONS: Even in the era of novel agent induction therapies, in our series of newly diagnosed 23 patients with a gain of 1q21, the failure rate of induction was 30% with a median PFS of 14 months. Moreover, these patients had a particularly aggressive clinical course with both medullary and extramedullary plasmacytomas, suggesting that PET-CTs, rather than just skeletal surveys should be considered for initial staging and monitoring. Also, given an unusually high incidence (13%) of early onset CNS disease, prompt CSF evaluation and brain MRI should be performed on patients with neurologic symptoms or signs. We recommend that gain or amplification of q21 identified either by FISH (even without CD138 selection – as demonstrated here) or GEP be prospectively studied in patients with newly diagnosed MM with consideration of novel therapeutic approaches. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Accumulation of CD69+ Terminal Effector CD8+ T cells occurs in the bone marrow of newly diagnosed Myeloma patients who lack protective clonal Vb expanded cytotoxic T cells

2019 ◽  
Vol 19 (10) ◽  
pp. e29
Author(s):  
Christian Bryant ◽  
Ka Hei Aleks Lau ◽  
Slavica Vuckovic ◽  
Felix Marsh-Wakefield ◽  
Annabel Kruzins ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Cd8 T Cells ◽  
Cytotoxic T Cells ◽  
Newly Diagnosed ◽  
Terminal Effector

scholarly journals Single-Cell Deconvolution Identifies T-Cell Correlates of Response to PD-1 Blockade Treatment in AML

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1950-1950
Author(s):  
Xingyue An ◽  
Jay R Adolacion ◽  
Mansour Alfayez ◽  
Jairo Matthews ◽  
Wilmer Flores ◽  
...  
Keyword(s):  
Gene Expression ◽  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Response Rate ◽  
Patient Stratification ◽  
Rna Seq ◽  
Cell Library

Background The combination of the αPD-1 (nivolumab) and hypomethylating agent azacytidine demonstrated encouraging response rate and overall survival in relapsed/refractory acute myeloid leukemia (AML) patients, compared to azacytidine alone1 (NCT02397720). However, the percentage of the patient who achieved an IWG 2016 response to such therapy was still limited (overall response rate = 33%), so it is desirable to have early predictive biomarkers to facilitate patient stratification and selection for future trials. A better understanding of T cells (primary targets of αPD-1 therapy) from bone marrow (BM) and peripheral blood (PB) in AML pre-therapy and on-therapy should yield valuable insights on the treatment-induced anti-tumor response. Methods We performed whole transcriptomic profiling (RNA-sequencing, RNA-seq) on T cells from a cohort of AML patients who were enrolled in the clinical trial mentioned above, and treated with azacytidine and nivolumab (Table 1). Sixty-four FACS-sorted patient-derived T cell samples from cryopreserved peripheral blood (PB) or bone marrow (BM) aspirates, which were collected pre-therapy (T0) and after the first round of treatment (end of cycle one, EOC1), were evaluated. By leveraging subset definitions based on scRNA-seq results from T cells of cancer patients, we implemented deconvolution of our bulk T-cell RNA-seq data to obtain the relative abundance of different T-cell subsets (in-silico dissection without physical isolation). Results We performed ex vivo cDNA library preparation on 2,000-100,000 sorted T cells and yielded a minimal of 17 million sequencing reads per BM T-cell library and 2.6 million sequencing reads per PBMC T-cell library. We compared the gene expression profile of peripheral blood T cells from AML patients (CD4: n = 16; CD8: n = 15) and healthy donors (HD, n = 8)2,3 to validate our methodology. The deconvolution results were consistent with previously published flow-cytometry data profiling cancer patients4,5 (Figure 1). In comparison with HDs, circulating CD4 T cells from AML patients consisted of a higher frequency of Treg (AML versus HD: 5.1% versus 0.6%, P<0.0001)4. Peripheral blood CD8 T cells from AML patients were with a significantly lower frequency of naïve (AML versus HD: 22.1% versus 55.9%, P<0.0001), a higher frequency of effector phenotype (AML versus HD: 35.6% versus 6.7%, P =0.0011), and an increasing frequency of exhausted phenotype (AML versus HD: 19.1% versus 0.0%, P<0.0001)5. Independent of the clinical responses attained on the azacytidine and nivolumab, comparison of the pre-therapy CD8 T cells from BM and PB from all AML patients using both gene set enrichment analysis and deconvolution indicated that the BM CD8 T cells were more activated/differentiated compared with PBMC CD8 T cells, likely reflective of an ongoing immune response against the AML. We also found treatment-induced gene expression profile changes in the AML circulating CD8 T cells, characterized by increased cell metabolism and cell proliferation. Deconvolution identified that pre-therapy relative abundance of exhausted (CD3+CD8+PD-1+CD45RO+) and effector (CD3+CD8+CD45RA+Tbet+PD-1lo) CD8 T cells (plasticity) could serve as subpopulations relevant for patient stratification (Figure 2). We further validated these results using CyTOF wherein these same subpopulations were differentially abundant between responders and non-responders at the pre-therapy time-point. Conclusions Collectively, the analyses of RNA-seq of CD8 T cells from AML patients on the azacytidine with nivolumab trial revealed that (1) the PD-1 blockade-based treatment-induced gene expression profiling changes (increased cell proliferation and metabolism) of CD8 T cells are detectable as early as EOC1 in the peripheral blood; (2) specific subpopulations of plastic CD8 T cells identified may have the potential to serve as an actionable biomarker to select AML patients most likely to benefit from such immune checkpoint therapies. These findings need to be confirmed in larger studies, planned or currently being conducted, with αPD-1 based therapies in AML and MDS. References 1. Daver, N. et al.Cancer Discov.9, 370-383 (2019). 2. Corces, M. R. et al.Nat. Genet.48, 1193-1203 (2016). 3. Linsley, P. S. et al. PLoS One9, (2014). 4. Szczepanski, M. J. et al.Clin. Cancer Res.15, 3325-3332 (2009). 5. Knaus, H. A. et al.JCI Insight3, (2018). Disclosures Varadarajan: CellChorus: Employment, Equity Ownership.

Changes of Intracellular Signal Pathway of mTOR/S6 in T Cells of Refractory/Relapsed Aplastic Anemia

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 4110-4110
Author(s):  
Guangsheng He ◽  
Xiang Zhang ◽  
Wu Depei ◽  
Mingqing Zhu ◽  
Aining Sun
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Aplastic Anemia ◽  
Cd8 T Cells ◽  
Signal Pathway ◽  
Deficiency Anemia ◽  
Newly Diagnosed ◽  
Ifn Γ ◽  
Intracellular Signal ◽  
Normal Controls

Abstract Objects: To explore the activation state of intracellular signal pathway of mTOR/S6 in refractory/relapsed aplastic anemia (AA); the effect of rapamycin (RAPA) and CTLA-4 immunoglobulin (CTLA-4Ig) on this signal pathway in refractory/relapsed AA. Methods: Samples were collected from 13 refractory/relapsed AA patients [male: 6, female: 7, media age: 27 years (from 7 to 66years)], 8 newly diagnosed patients with severe aplastic anemia (SAA) [male: 5, female: 3, media age: 21.5 years (from 12 to 58 years)]. The intracellular percentages of p-mTOR, p-S6 and IFN-γ of CD3+, CD3+CD8+ and CD3+CD8− T cells in bone marrow were detected by flow cytometry (FCM). 10 iron deficiency anemia (IDA) patients [male: 3, female: 7, media age: 44 years (from 31 to 72 years)] were determined as case controls and normal controls respectively. After exposure to RAPA and CTLA-4Ig respectively, samples were detected by FCM for the expression of p-mTOR, p-S6 and IFN-γ in CD3+, CD3+CD8+ and CD3+CD8− T cells in bone marrow, in order to estimate the effect of RAPA and CTLA-4Ig on the pathway of mTOR/S6 in refractory/relapsed AA. Results: In refractory/relapsed AA, measurement of p-mTOR, p-S6 and IFN-γ in CD3+, CD3+CD8− and CD3+CD8+ T cells were (42.42±26.44)%, (44.38±24.95)%, (51.89±27.00)%; (6.47±2.72)%, (9.16±2.89)%, (9.61±5.34)%; (18.87±10.05)%, (13.17±5.88)%, (20.07±15.16)%, respectively; and showed an increased level compared to normal controls [(1.54±1.51)%; P=0.000], [(1.94±1.08)%; P=0.000], [(2.04±2.03)%; P=0.000]; [(0.83±0.82)%; P=0.000], [(0.91±0.88)%; P=0.000], [(0.95±0.93)%; P=0.000]; [(4.42±3.55)%; P=0.000], [(2.35±1.69)%; P=0.000], [(4.73±4.43)%; P=0.004]. In newly diagnosed patients with SAA, the levels of p-mTOR and p-S6 in CD3+, CD3+CD8− and CD3+CD8+ T cells were (1.71±1.66)%, (2.28±2.15)%, (1.59±1.52)%; (1.23±1.13)%, (1.23±1.07)%, (1.76±1.68)% respectively, and they were similar to normal controls (P&gt;0.05), but significantly lower than those of refractory/relapsed AA (P&lt;0.01). Expression of IFN-γ in CD3+, CD3+CD8− and CD3+CD8+ T cells was higher than normal controls with (10.38±3.83)%, (6.11±1.91)%, (13.14±7.05)% (P&lt;0.01). The percentages of CD3+IFN-γ+ and CD3+CD8−IFN-γ+ were lower than refractory/relapsed AA (P&lt;0.05), while it was comparable for CD3+CD8+IFN-γ+ cells between the two groups (P&gt;0.05). Exposed to RAPA, the expression of p-mTOR, p-S6 and IFN-γ in CD3+, CD3+CD8− and CD3+CD8+ T cells decreased markedly (P&lt;0.05) to (12.44±12.41)% (12.60±12.57)%, (16.85±15.64)%; (1.49±1.45)%, (1.46±1.43)%, (1.55±1.54)%; and (4.29±4.23)%, (3.16±3.32)%, (10.70±10.63)% in refractory/relapsed AA. And treated with CTLA-4Ig could also cause a significant reduction of p-mTOR, p-S6 and IFN-γ in CD3+, CD3+CD8− and CD3+CD8+ T cells, which were (6.40±6.13)%, (8.32±7.76)%, (7.18±7.02)%; (1.08±1.08)%, (2.69±2.37)%, (1.60±1.56)%; (1.67±1.60)%, (2.39±2.12)%, (1.30±1.30)%, respectively, (P&lt;0.01). Conclusions: In refractory/relapsed AA, the signal pathway of mTOR/S6 was activated, and it was quiescent in normal controls and newly diagnosed patients with SAA. The expression of p-mTOR, p-S6 and IFN-γ of this signal pathway in refractory/relapsed AA could be suppressed by RAPA or CTLA-4Ig. The signal pathway of CD28/mTOR/S6/IFN-γ might take part in immune pathogenesis of refractory/relapsed AA, and was sensitive to RAPA and CTLA-4Ig. It was worth exploring the clinical values of the two drugs.

scholarly journals Bone marrow CD8 T cells express high frequency of PD-1 and exhibit reduced anti-leukemia response in newly diagnosed AML patients

2018 ◽  
Vol 8 (3) ◽  
Author(s):  
Bei Jia ◽  
Liru Wang ◽  
David F. Claxton ◽  
W Christopher Ehmann ◽  
Witold B. Rybka ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Cd8 T Cells ◽  
High Frequency ◽  
Newly Diagnosed

507 High dimensional flow cytometry analysis in newly diagnosed acute myeloid leukemia predicts patients outcomes

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A543-A543
Author(s):  
Francesco Mazziotta ◽  
Rupkatha Mukhopadhyay ◽  
Hanna A Knaus ◽  
Anish Chowdhury ◽  
Amanda Blackford ◽  
...  
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Myeloid Leukemia ◽  
Cd8 T Cell ◽  
Effector Memory ◽  
Newly Diagnosed ◽  
Response To Chemotherapy ◽  
Acute Myeloid

BackgroundWe have previously characterized phenotypic and transcriptional profile of CD8+ T cells in acute myeloid leukemia (AML) and their differences between responders vs. nonresponders to chemotherapy.1 Goal of ongoing work was to further probe uniqueness of AML in sculpting CD8+ T cell responses and the plasticity of their signatures upon chemotherapy response.MethodsWe first examined the cumulative expression of multiple inhibitory receptors (IRs) (detected by 2 different panels) on CD8+ T cells and created an IR-score which summarizes the relative amount of PD-1, Tim3, KLRG1, 2B4, CD160, CD57, and BTLA-positive CD8+ T-cells in relation to the well-characterized maturation states of CD8+ T cells. Serial bone marrow samples from 33 newly diagnosed AML patients with well-annotated clinical data (21 complete responders (CR) and 12 nonresponders (NR) to chemotherapy) and 11 healthy controls (HC) were analyzed. FInally, using custom made R code, we performed dimensionality reduction, clustering, and pseudotime analysis.ResultsThe IR-score discriminated NR and CR (p = 3e-02, AUC 0.84) after treatment with CD57 and KLRG1 accounting for most of this difference (p = 2e-02, AUC = 0.79). Next we investigated CD8+ T cell populations that best correlated with response to chemotherapy. FlowSOM revealed seven major clusters: naive and naive-like, CD28+KLRG1+ activated-effector, CD28+KLRG1+PD1+ dysfunctional, PD1+CD57+ senescent effector-memory and two clusters of terminally differentiated CD45RA+KLRG1+ cells. Since the activation and differentiation states accounted for most of the subpopulation variability, we grouped the clusters into resting (naive, naive-like), activated (activated-effector, dysfunctional), and terminally differentiated cells (senescent effector-memory, terminally differentiated). UMAP, developmental trajectories and differential abundance testing showed increased frequency of activated cells at diagnosis (p-adj = 2.9e-05) and of resting cells after treatment (p-adj = 1.3e-02) in CR, while terminally differentiated T cells prevailed in NR (p-adj = 5.3e-08) after treatment (figures 1 and 2).Abstract 507 Figure 1UMAP embedding of T cells in CR, NR, at diagnosis (BM_DG) and after chemotherapy (BM_post), HC colored by T cell state (resting, activated, terminal differentiated), overlaid with a contour plotAbstract 507 Figure 2Boxplots showing the differential cluster abundance and adjusted p-values for CR, NR, at diagnosis (BM_DG) and after chemotherapy (BM_post), HC in the three different T cell states (resting, activated, terminal differentiated)ConclusionsThe increased number of functional activated T cells at diagnosis and the persistence of a naive/naive-like reservoir at the time of response is a signature associated with achievement of CR. Lack of response (NR) correlates with accumulation of the terminally differentiated and senescent cells in the bone marrow. These results uncover an intertwined relationship between skewing of T cell differentiation and clinical response to chemotherapy. The data provide rationale to either remove senescent or augment activity of naïve/naïve-like T cells as a strategy to reinforce antileukemia immunity.ReferenceKnaus HA, Berglund S, Hackl H, et al. Signatures of CD8+ T cell dysfunction in AML patients and their reversibility with response to chemotherapy. JCI Insight 2018; 3(21).

scholarly journals Increased frequency of TIGIT+CD73-CD8+ T cells with a TOX+ TCF-1low profile in patients with newly diagnosed and relapsed AML

2021 ◽  
Vol 10 (1) ◽  
pp. 1930391
Author(s):  
F. Brauneck ◽  
F Haag ◽  
R. Woost ◽  
N. Wildner ◽  
E. Tolosa ◽  
...  
Keyword(s):  
T Cells ◽  
Cd8 T Cells ◽  
Newly Diagnosed

scholarly journals AB0041 CD8+ T CELLS HAVE AN ELEVATED PROLIFERATIVE CAPACITY IN GIANT CELL ARTERITIS

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 1323.1-1323
Author(s):  
R. Reitsema ◽  
R. Hid Cadena ◽  
W. Abdulahad ◽  
A. Boots ◽  
P. Heeringa ◽  
...  
Keyword(s):  
T Cells ◽  
Giant Cell Arteritis ◽  
Giant Cell ◽  
Cd8 T Cells ◽  
Mononuclear Cells ◽  
Effector Memory ◽  
Newly Diagnosed ◽  
Proliferative Capacity ◽  
Ki 67 ◽  
Age And Sex

Background:Giant cell arteritis (GCA) is the most frequent form of systemic vasculitis affecting the large- and medium-sized vessels. The involvement of innate immune cells and CD4+ T cells in the pathogenesis of GCA has been extensively studied. Interestingly, recent findings suggest a role for CD8+ T cells in disease development (1). However, CD8+ subsets and their functional capacities have not yet been studied in detail.Objectives:This study aims to characterize the phenotype and proliferative capacity of CD8+ T cells in newly diagnosed GCA patients and GCA patients in remission compared to healthy age- and sex- matched controls.Methods:To determine the phenotype of CD8+ T cells in GCA, newly diagnosed, untreated GCA patients (baseline, n=14), GCA patients in stable glucocorticoid-free remission (GC-FR, n=10) and age- and sex-matched healthy controls (HCs, n=18) were enrolled. Peripheral blood mononuclear cells (PBMCs) were stained with fluorochrome-conjugated antibodies directed against CD3, CD4, CD8, CCR7, CD45RO, Ki-67, CD69 and CD25 and analyzed by flow cytometry. The following differentiation subsets were defined: CD8+ T naive (CD45RO-CCR7+), central memory (TCM, CD45RO+CCR7+), effector memory (TEM, CD45RO+CCR7-) and effector memory re-expressing CD45RA (TEMRA, CD45RO-CCR7-) cells. Secondly, the proliferative capacity of CD8+ T cells was determined in isolated CD3+ T cells of 10 GCA baseline, 10 GCA GC-FR patients and 19 HCs after 5 days of stimulation with plate-bound anti-CD3 or anti-CD3 plus soluble anti-CD28 using a dye-based proliferation assay.Results:A reduced frequency of CD8+ TEMcells was found in GCA baseline patients compared to HCs (p=0.025). Furthermore, a higher frequency of Ki-67+ cells was detected among CD8+ TEMcells in GCA baseline patients than in HCs (p=0.0007), suggesting a higher proliferative activityin vivo.In addition,in vitrostimulation with anti-CD3 and anti-CD3+anti-CD28 led to higher percentages of divided CD8+ T cells in GCA baseline and GC-FR patients than in HCs (p<0.05). Moreover, the frequencies of CD8+ TEMRAcells and the percentage of divided CD8+ T cells upon CD3 stimulation strongly correlated in GCA baseline patients (R=0.79, p=0.009) and GCA GC-FR patients (R=0.67, p=0.039) but not in HCs (R=0.31, p=0.25).Conclusion:GCA baseline patients demonstrate a higher frequency of proliferating circulating CD8+ TEMcells, defined by Ki-67 expression, than HCs. In addition, functional data on induced proliferative capacity suggest that CD8+ T cells from GCA baseline patients are more rapidly activated by crosslinking CD3 and CD3+CD28, suggesting either reduced regulation in these patients or more intrinsic threshold changes. Furthermore, the induced proliferative capacity is also elevated in patients in stable glucocorticoid-free remission. Whether the increased proliferative capacity of total CD8+ T cells in GCA patients is causally linked to the increased frequencies of CD8+ TEMRAcells in these patients requires further investigation.References:[1]Samson M, Ly KH, Tournier B, Janikashvili N, Trad M, Ciudad M, et al. Involvement and prognosis value of CD8+ T cells in giant cell arteritis. J Autoimmun. 2016;72:73–83.Disclosure of Interests:Rosanne Reitsema: None declared, Rebeca Hid Cadena: None declared, Wayel Abdulahad: None declared, Annemieke Boots Consultant of: Grünenthal Gmbh until 2017, Peter Heeringa: None declared, Elisabeth Brouwer Consultant of: Roche (consultancy fee 2017 and 2018 paid to the UMCG), Speakers bureau: Roche (2017 and 2018 paid to the UMCG)

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