Purpose: Gastrin-releasing peptide (GRP) is a growth factor for small cell lung cancer (SCLC). GRP belongs to the bombesin peptide
family and has significant homology to bombesin. We constructed a ...bispecific molecule, OKT3xAntag2, by conjugating a monoclonal
antibody OKT3 (anti-CD3) with a bombesin/GRP antagonist (Antag2) and evaluated cytotoxicity against SCLC cells.
Experimental Design: We tested binding of the bispecific molecule to SCLC cell lines and T cells by flow cytometry, antibody-dependent cellular
cytotoxicity (ADCC) of SCLC cells in vitro and in a murine SCLC xenograft model. We studied SCLC apoptosis and necrosis during ADCC and the activity and cleavage of
caspase-3, caspase-9, and poly(ADP-ribose) polymerase (PARP).
Results: The bispecific molecule functions as a cross-linker between T cells and SCLC cells, induces T cell activation, and mediates
ADCC of SCLC cells; 40% to 80% growth inhibition of SCLC cells mediated by the bispecific molecule at low effector to target
cell ratios was achieved. Activation of T cells by the bispecific molecule resulted in significant increases in IFNγ production
and apoptosis and necrosis of SCLC cells associated with cleavage of PARP and caspase-3. Targeted immunotherapy with the bispecific
molecule–armed human T cells significantly reduced SCLC tumor burdens in a mouse model.
Conclusion: The bispecific molecule OKT3xAntag2 mediates growth inhibition and apoptosis of SCLC cells by activated T cells through activation
and cleavage of caspase-3 and PARP in vitro and in vivo . Clinical trials of this bispecific molecule through adoptive transfer of ex vivo activated T cells in GRP receptor–positive tumors, such as SCLC, are warranted.
This study investigated whether TNF-α, Toll-like receptors (TLRs) 7/8 agonist resiquimod (R848), the TLR4 agonist lipopolysaccharide (LPS) and their combinations can enhance autologous AML-reactive T ...cell generation in an in vitro culture. AML peripheral blood or bone marrow mononuclear cells were cultured in medium supplemented with GM-CSF/IL-4 to induce dendritic cell (DC) differentiation of AML blasts (AML-DC). The impact of TNF-α, LPS, R848 and their combinations on AML-DC cultures was analyzed. Significantly enhanced CD80, CD40, CD83, CD54, HLA-DR and CD86 expression of AML cells was observed by addition of TNF-α, LPS, R848 alone or combinations. Induced CD80 expression of AML cells was significantly higher through the combination of TNF-α, LPS and R848 (T + L + R) than that by T alone. CTL induced from T + L + R, T + R, T + L, L + R and R, but not T, L alone stimulated cultures showed significantly higher IFN-γ release than the medium control in response to autologous AML cells. IFN-γ release by T + L + R was significantly higher than T or L alone, and T + R was significantly higher than T alone. CTL generated from T + L + R, T + L, T + R, L + R and L alone exerted significantly higher AML cell killing than medium control. AML cell killing by T + L + R and T + R was significantly higher than T or R alone. These results indicate that the combination of T + L + R induces a significantly enhanced antigen presentation effect of AML-DC. We speculate that the complementary effects of reagent combinations may better address the heterogeneity of responses to any single agent in AML cells from different patients.
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EMUNI, GEOZS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UL, UM, UPUK, VKSCE, ZAGLJ
Methods that enable monitoring of therapeutic efficacy of autologous chimeric antigen receptor (CAR) T-cell therapy will be clinically useful. The aim of this study is to demonstrate the feasibility ...of blood-derived cell-free DNA (cfDNA) to predict CAR T-cell therapy response in patients with refractory B-cell lymphomas. Whole blood was collected before and throughout CAR T-cell therapy until day 154. Low-coverage (∼0.4×), genome-wide cfDNA sequencing, similar to that established for noninvasive prenatal testing, was performed. The genomic instability number (GIN) was used to quantify plasma copy number alteration level. Twelve patients were enrolled. Seven (58%) patients achieved a complete response (CR); 2 (25%), a partial response. Median progression-free survival was 99 days; median overall survival was not reached (median follow-up, 247 days). Altogether, 127 blood samples were analyzed (median, 10 samples/patient range 8-13). All 5 patients who remained in CR at the time of last measurement had GIN <170 (threshold). Two patients who attained CR, but later relapsed, and all but one patient who had best response other than CR had last GIN measurement of >170. In 5 of 6 patients with relapsed or progressive disease, increasing GIN was observed before the diagnosis by imaging. The abundance of CAR T-cell construct (absolute number of construct copies relative to the number of human genome equivalents) also showed a trend to correlate with outcome (day 10, P = .052). These data describe a proof-of-concept for the use of multiple liquid biopsy technologies to monitor therapeutic response in B-cell lymphoma patients receiving CAR T-cell therapy.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
BACKGROUND: Interpatient variability in the kinetics of peripheral blood progenitor cell (PBPC) mobilization is commonly seen with conventional chemotherapy‐based mobilization regimens. This ...necessitates the availability of leukapheresis (LP) facilities 7 days a week.
STUDY DESIGN AND METHODS: The efficacy of an approach where LP was invariably commenced on Day 11 after intermediate‐dose cyclophosphamide followed by sequential administration of granulocyte‐macrophage–colony‐stimulating factor (CSF) and granulocyte–CSF (Cy/GM/G) was retrospectively analyzed in 225 consecutive, unselected patients undergoing autologous hematopoietic stem cell transplantation for all diagnoses other than acute leukemia at our center. Cy/GM/G was scheduled to avoid weekend LP.
RESULTS: After Cy/GM/G, a CD34+ cell yield of at least 2.0 × 106 per kg was achieved in 90.7 percent of patients. Optimal yield (OY; ≥5 × 106 or 10 × 106 CD34+ cells/kg depending on diagnosis) was achieved in 67.6 percent of patients. Only three patients (1.3%) required LP on Saturday or Sunday. Febrile neutropenia (FN) was encountered in 5.3 percent. PBPC yield was highest on Day 1 of LP (p < 0.001). In multivariate analyses, platelet (PLT) count on Day 1 of LP (PLT‐D1LP) was positively associated with achievement of OY (p < 0.001). PLT‐D1LP and diagnosis of myeloma were associated with a shorter time to achieve a CD34+ cell yield of at least 5 × 106 per kg (p < 0.001 and p = 0.002, respectively).
CONCLUSION: Cy/GM/G with scheduled LP commencement on Day 11 enables optimal CD34+ cell yields in most patients undergoing autologous transplantation, despite a low risk of FN and avoidance of weekend LP.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Patients (pts) with relapsed hematologic malignancies (HM) after alloHCT are a unique population, given the potential to harness a dormant graft-vs.-tumor effect therapeutically. We previously ...reported that CTLA-4 blockade with ipilimumab was feasible and active in this population (Davids et al., N Eng J Med, 2016). In retrospective studies, anti-PD1 antibodies were active in pts with relapsed lymphoid malignancies after alloHCT, though with substantial toxcity due to GVHD (Herbaux et al. and Haverkos et al., Blood, 2017). Here, we report on the first prospective clinical trial of PD1 blockade in pts with relapsed HM after alloHCT.
The primary objectives in this ph I/Ib, multicenter, investigator-initiated, CTEP-sponsored study (CTEP 9204) were to determine MTD and evaluate safety of nivolumab (nivo). Secondary objectives were to assess efficacy and immunologic correlates. Pts with any HM with relapse or persistent disease after alloHCT were eligible. Nivo was initially given to a 1 mg/kg cohort, with planned escalation to a 3 mg/kg cohort or de-escalation to a 0.5 mg/kg cohort depending on toxicities. Nivo was dosed q2 wks until progression or unacceptable toxicity, and disease-specific response evaluations were q4 cycles.
A total of 28 pts with relapsed HM after alloHCT were treated. Median age was 57 (range 27-76), and pts had the following HM: AML (n=11), MDS (n=7), Hodgkin lymphoma (HL, n=5), non-Hodgkin lymphoma (NHL, n=3), MPD and CLL (n=1 each). Median number of prior therapies was 2 (range 1-9), and 18/28 (64%) had progressed after at least 1 prior therapy for relapse post alloHCT. The median time from alloHCT to study enrollment was 21 mo. (range 5.7-174 mo.).
Six pts were treated initially with nivo 1 mg/kg. Two immune-related adverse events (irAEs) resulted in DLTs, including one pt with sepsis and fatal ARDS, and one pt with new anti-phospholipid antibodies and a fatal thrombotic cerebral vascular accident. Other irAEs included gr3 pneumonitis and transaminitis (n=1 each). One pt had cGVHD (NIH mild). Response was observed in 3/6 pts, including 1 CR (PMBCL) and 2 PR (HL and CMML).
Due to the toxicities at 1 mg/kg, a cohort of 8 pts was then treated with nivo 0.5 mg/kg, which was generally well-tolerated, with no DLTs. A phase Ib expansion cohort then accrued 14 more pts at 0.5 mg/kg. Accrual was terminated after 14 pts were treated due to meeting the protocol-defined stopping rule of ≥4 DLTs in the first 15 pts in this cohort. These DLTs included 2 cases of grade III acute GVHD (liver and gut) as well as gr3 elevated bilirubin (n=1) and gr3 transaminitis (n=1) which did not recover to ≤gr1 within 4 wks. The 2 pts with liver dysfunction without histological evidence of GVHD eventually improved, but both pts with GVHD died due to complications from GVHD. Other toxicities included gr4 lipase elevation, gr3 rash, gr3 transaminitis, gr3 orthostatic hypotension, and gr2 seizure in a pt with a known seizure disorder (n=1 each). In the 22 pts treated at nivo 0.5 mg/kg, 10 pts (45%) had new onset or worsening of GVHD, including 1 with aGVHD only, 7 with cGVHD only (3 of whom had baseline cGVHD), and 2 with both acute and cGVHD. Shorter time from alloHCT was significantly associated with higher risk of developing GVHD (p=0.019). The overall response rate in the 19 evaluable pts treated at nivo 0.5 mg/kg was 16%, including 1 pt with HL with CR and 1 pt each with HL and AML achieving PR. Nine pts had stable disease for at least 1 response evaluation, and 7 pts had progressive disease as best response.
Studywide, the overall response rate was 24% (6/25), the median number of cycles received was 3 (range 1-25), and 12/28 (43%) had at least 1 dose delay due to toxicity. With a median follow-up of 3.9 mo. (range 1.4-20.9 mo.), the 6 mo. PFS and OS were 39% and 61%, respectively.
In this first prospective clinical trial of an anti-PD1 antibody for relapsed HM post-alloHCT, severe GVHD and irAEs occurred, even at the lower dose of nivo 0.5 mg/kg, leading to early closure due to toxicity. Modest anti-tumor activity was observed mainly in lymphoid malignancies known already to be responsive to anti-PD1 therapy, which may justify further exploration of anti-PD1 therapy in those populations in trials with strategies to mitigate toxicity; however, given the more favorable safety and efficacy profile of anti-CTLA-4 therapy in other HM, our future studies focus on combining ipilimumab with novel partners to improve outcomes.
Davids:Celgene: Consultancy; MEI Pharma: Consultancy, Research Funding; Verastem: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; TG Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Astra-Zeneca: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Roche/Genentech: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pharmacyclics: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Merck: Consultancy; BMS: Research Funding; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; AbbVie, Inc: Consultancy, Membership on an entity's Board of Directors or advisory committees; Surface Oncology: Research Funding. Costello:Celgene: Consultancy; Takeda: Consultancy; Poseida Therapeutics, Inc.: Research Funding. Herrera:Seattle Genetics: Research Funding; Pharmacyclics: Consultancy, Research Funding; Merck, Inc.: Consultancy, Research Funding; KiTE Pharma: Consultancy, Research Funding; Immune Design: Research Funding; AstraZeneca: Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Gilead Sciences: Research Funding. Locke:Novartis Pharmaceuticals: Other: Scientific Advisor; Cellular BioMedicine Group Inc.: Consultancy; Kite Pharma: Other: Scientific Advisor. Chen:Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; Magenta Therapeutics: Consultancy; REGiMMUNE: Consultancy; Takeda Pharmaceuticals: Consultancy. Nikiforow:Kite Pharma: Consultancy. Ho:Jazz Pharmaceuticals: Consultancy. Wu:Neon Therapeutics: Equity Ownership. Soiffer:Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
A key aim of systems biology is the reconstruction of molecular networks. We do not yet, however, have networks that integrate information from all datasets available for a particular clinical ...condition. This is in part due to the limited scalability, in terms of required computational time and power, of existing algorithms. Network reconstruction methods should also be scalable in the sense of allowing scientists from different backgrounds to efficiently integrate additional data. We present a network model of acute myeloid leukemia (AML). In the current version (AML 2.1), we have used gene expression data (both microarray and RNA-seq) from 5 different studies comprising a total of 771 AML samples and a protein-protein interactions dataset. Our scalable network reconstruction method is in part based on the well-known property of gene expression correlation among interacting molecules. The difficulty of distinguishing between direct and indirect interactions is addressed by optimizing the coefficient of variation of gene expression, using a validated gold-standard dataset of direct interactions. Computational time is much reduced compared to other network reconstruction methods. A key feature is the study of the reproducibility of interactions found in independent clinical datasets. An analysis of the most significant clusters, and of the network properties (intraset efficiency, degree, betweenness centrality, and PageRank) of common AML mutations demonstrated the biological significance of the network. A statistical analysis of the response of blast cells from 11 AML patients to a library of kinase inhibitors provided an experimental validation of the network. A combination of network and experimental data identified CDK1, CDK2, CDK4, and CDK6 and other kinases as potential therapeutic targets in AML.
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