Cytotoxic T-lymphocyte Antigen-4 Blockade Research Articles

CTLA-4 Blockade of Natural Killer Cells Increases Cytotoxicity against Acute Lymphoid Leukaemia Cells Neda.

There is interest in using cytotoxic T lymphocyte antigen-4 (CTLA-4) immunotherapy to treat blood cancers. Unfortunately, patients with acute lymphoblastic leukaemia (ALL) frequently exhibit resistance to treatment and natural killer (NK) cell exhaustion. This study aims to increase the cytotoxic potency of natural killer cells by using CTLA-4 to block the Nalm-6 leukaemia cell line. In this experimental study, NK cells were purified from the peripheral blood mononuclear cells (PBMCs) of 10 healthy people and assessed by flow cytometry for purity and viability. The purified cells were activated overnight at 37°C and 5% CO2 with interleukin-15 (IL-15, 10 ng/ml) followed by evaluation of expressions of CTLA-4, activating and inhibitory receptors, and the release of interferon gamma (IFN-γ) and granzyme B (GZM B). CTLA-4 expression on NK cells from recurrent ALL patients was also evaluated. Finally, the cytotoxic activity of NK cells was assessed after the CTLA-4 blockade. The purity of the isolated cells was 96.58 ± 2.57%. Isolated NK cells activated with IL-15 resulted in significantly higher CTLA-4 expression (8.75%, P<0.05). Similarly, CTLA-4 expression on the surface of NK cells from patients with ALL was higher (7.46%) compared to healthy individuals (1.46%, P<0.05). IL-15 reduced NKG2A expression (P<0.01), and increased expressions of NKP30 (P<0.05) and NKP46 (P<0.01). The activated NK cells released more IFN-γ (P<0.5) and GZM B (P<0.01) compared to unactivated NK cells. Blockade of CTLA-4 enhanced the NK cell killing potential against Nalm-6 cells (56.3%, P<0.05); however, IFN-γ and GZM B levels were not statistically different between the blocked and non-blocked groups. Our findings suggest that CTLA-4 blockage of Nalm-6 cells causes an increase in antitumour activity of NK cells against these cells. Our study also provides evidence for the potential of cancer immunotherapy treatment using blocking anti-CTLA-4 mAbs.

Granulomatous and Sarcoid-like Immune-Related Adverse Events following CTLA4 and PD1 Blockade Adjuvant Therapy of Melanoma: A Combined Analysis of ECOG-ACRIN E1609 and SWOG S1404 Phase III Trials and a Literature Review.

Treatment with immune checkpoint inhibitors (ICIs) has been linked to granulomatous and sarcoid-like lesions (GSLs) affecting different organs. This study sought to evaluate GSL incidence in patients with high-risk melanoma treated with cytotoxic T-lymphocyte antigen 4 (CTLA4) or programmed cell death 1 (PD1) blockade adjuvant therapy in two clinical trials: ECOG-ACRIN E1609 and SWOG S1404. Descriptions and GSL severity ratings were recorded. Data were collected from ECOG-ACRIN E1609 and SWOG S1404. Descriptive statistics along with GSL severity grades were reported. Additionally, a literature review for such cases was summarized. A total of 11 GSL cases were reported among 2878 patients treated with either ICI or with High-Dose Interferon Alfa-2b (HDI) in ECOG-ACRIN E1609 and SWOG S1404 trials. Cases were numerically more commonly reported with ipi10, followed by pembrolizumab, ipi3, and HDI, respectively. Most of the cases were grade III. Further, organs involved included lung, mediastinal lymph nodes, skin and subcutaneous tissue, and eye. Furthermore, a summary of 62 reports in the literature was described. GSLs following anti-CTLA4 and anti-PD1 antibody therapy in patients with melanoma were reported unusually. Reported cases ranged in grade from I to III and appeared manageable. Careful attention to these events and their reporting will be essential to better guide practice and management guidelines.

Elaborating on anti CTLA-4 mechanisms of action using an agent-based modeling approach

Anti CTLA-4 therapy is aimed at blocking the Cytotoxic T-lymphocyte antigen-4 (CTLA-4), a key cancer immunity cycle checkpoint. The mechanism of action of CTLA-4 may be described as a dynamic competition for the B7 ligand which, subsequently, interferes with the CD28-B7 costimulatory pathway. Anti CTLA-4 blockade enhances the process of cognate T cell activation and leads to a broadening of the T cell repertoire. In the present work, we used an agent-based modeling (ABM) platform of T cell immune response development, to explore hypothetical modes of anti CTLA-4 action. The model features a selected number of activated T cell clones, calculated based on combined random and chemotactically-driven encounters with antigen-presenting dendritic cells (DCs) and a distribution of individual T cell affinities to the antigen of interest. The proposed model can be used as a quantitative tool to explore various hypotheses on T cell immunity regulation and validate these against experimental data. A comprehensive ABM model analysis of immune response dynamic simulations revealed several putative anti CTLA-4 mechanisms of action, including: (i) an increase in the probability of primary activation of lymphocytes; (ii) T cell activation enhancement via a prolongation of short contacts with dendritic cells; and (iii) an increase in the maximum level of activation signal (or saturation), accumulated through a series of short contacts with DCs. The modeling work further demonstrates that it is only when considering jointly these various modes of anti CTLA-4 effects on the T cell immune response dynamics that a biologically meaningful increase in both the production of activated cells and the expansion of the T cell repertoire is observed. These model-based results are overall consistent with the collective biological knowledge on the functional role of CTLA-4. Furthermore, the ABM presented here may allow to interrogate various mechanistic scenarios underlying adverse events mediated by anti CTLA-4 pharmacologic therapies.

The Role of Cytotoxic T-Lymphocyte Antigen 4 in the Pathogenesis of Multiple Sclerosis.

Multiple sclerosis (MS) is an autoimmune neurodegenerative disorder of the central nervous system that presents heterogeneous clinical manifestations and course. It has been shown that different immune checkpoints, including Cytotoxic T-Lymphocyte Antigen 4 (CTLA-4), can be involved in the pathogenesis of MS. CTLA-4 is a critical regulator of T-cell homeostasis and self-tolerance and represents a key inhibitor of autoimmunity. In this scopingreview, we resume the current preclinical and clinical studies investigating the role of CTLA-4 in MS with different approaches. While some of these studies assessed the expression levels of CTLA-4 on T cells by comparing MS patients with healthy controls, others focused on the evaluation of the effects of common MS therapies on CTLA-4 modulation or on the study of the CTLA-4 blockade or deficiency in experimental autoimmune encephalomyelitis models. Moreover, other studies in this field aimed to discover if the CTLA-4 gene might be involved in the predisposition to MS, whereas others evaluated the effects of treatment with CTLA4-Ig in MS. Although these results are of great interest, they are often conflicting. Therefore, further studies are needed to reveal the exact mechanisms underlying the action of a crucial immune checkpoint such as CTLA-4 in MS to identify novel immunotherapeutic strategies for MS patients.

CD80 Expression on Tumor Cells Alters Tumor Microenvironment and Efficacy of Cancer Immunotherapy by CTLA-4 Blockade.

Simple SummaryThe recent discovery of immune checkpoint inhibitors constituted a breakthrough in cancer treatment, but most patients are resistant to this therapy. Although the co-stimulatory molecule cluster of differentiation (CD) 80 has been detected in several types of tumor cells, its role in the tumor microenvironment and its sensitivity to immune checkpoint blockade are unclear. We, therefore, introduced a clinically relevant mouse tumor model with deactivated CD80. The deactivation promoted a “hot” tumor microenvironment and enhanced the sensitivity to immune checkpoint blockade with antibody against the cytotoxic T-lymphocyte antigen 4 (CTLA-4). This study contributed to the research into predictive markers to select patients who are suitable for immune checkpoint blockade therapy and suggested the development of a novel cancer immunotherapy based on a tumor-cell-targeted CD80 blockade. Cluster of differentiation (CD) 80 is mainly expressed in immune cells but can also be found in several types of cancer cells. This molecule may either activate or inhibit immune reactions. Here, we determined the immunosuppressive role of CD80 in the tumor microenvironment by CRISPR/Cas9-mediated deactivation of the corresponding gene in the mouse oncogenic TC-1 cell line. The tumor cells with deactivated CD80 (TC-1/dCD80-1) were more immunogenic than parental cells and induced tumors that gained sensitivity to cytotoxic T-lymphocyte antigen 4 (CTLA-4) blockade, as compared with the TC-1 cells. In vivo depletion experiments showed that the deactivation of CD80 switched the pro-tumorigenic effect of macrophages observed in TC-1-induced tumors into an anti-tumorigenic effect in TC-1/dCD80-1 tumors and induced the pro-tumorigenic activity of CD4+ cells. Moreover, the frequency of lymphoid and myeloid cells and the CTLA-4 expression by T helper (Th)17 cells were increased in TC-1/dCD80-1- compared with that in the TC-1-induced tumors. CTLA-4 blockade downregulated the frequencies of most immune cell types and upregulated the frequency of M2 macrophages in the TC-1 tumors, while it increased the frequency of lymphoid cells in TC-1/dCD80-1-induced tumors. Furthermore, the anti-CTLA-4 therapy enhanced the frequency of CD8+ T cells as well as CD4+ T cells, especially for a Th1 subset. Regulatory T cells (Treg) formed the most abundant CD4+ T cell subset in untreated tumors. The anti-CTLA-4 treatment downregulated the frequency of Treg cells with limited immunosuppressive potential in the TC-1 tumors, whereas it enriched this type of Treg cells and decreased the Treg cells with high immunosuppressive potential in TC-1/dCD80-1-induced tumors. The immunosuppressive role of tumor-cell-expressed CD80 should be considered in research into biomarkers for the prediction of cancer patients’ sensitivity to immune checkpoint inhibitors and for the development of a tumor-cell-specific CD80 blockade.

Immune Co-inhibitory Receptors PD-1, CTLA-4, TIM-3, LAG-3, and TIGIT in Medullary Thyroid Cancers: A Large Cohort Study.

Programmed cell death protein-1 (PD-1), cytotoxic T-lymphocyte antigen 4 (CTLA-4), T-cell immunoglobulin and mucin-domain containing-3 (TIM-3), lymphocyte activation gene-3 (LAG-3), and T-cell immunoglobulin and ITIM domain (TIGIT) are considered major immune co-inhibitory receptors (CIRs) and the most promising immunotherapeutic targets in cancer treatment, but they are largely unexplored in medullary thyroid carcinoma (MTC). We aimed to provide the first evidence regarding the expression profiles and clinical significance of CIRs in a large cohort of MTC patients. In total, 200 MTC patients who received initial surgery in our hospital were included. Immunohistochemistry was performed to evaluate CIR expressions in tissue microarrays (TMAs). Combined with the results of our previous programmed cell death ligand-1 (PD-L1) study, clinicopathologic and prognostic correlations of these proteins were retrospectively analyzed. TIM-3, PD-1, CTLA-4, LAG-3, and TIGIT positivity was detected in 96 (48.0%), 27 (13.5%), 25 (12.5%), 6 (3.0%), and 6 (3.0%) patients, respectively, in whom TIM-3, PD-1, and CTLA-4 expressions were positively correlated. Log-rank tests and multivariate Cox analyses both indicated that TIM-3, CTLA-4 expression, and PD-1/PD-L1 coexpression were associated with worse structural recurrence-free survival. In addition, among 20 patients who developed advanced disease during follow-up, 12 (60%) showed TIM-3 positivity, among whom 6 cases also had concurrent moderate to strong PD-1, PD-L1, or CTLA-4 expression. Using the currently largest TMA cohort of this rare cancer, we delineated the CIR expression profiles in MTC, and identified TIM-3, CTLA-4 expression, and PD-1/PD-L1 coexpression as promising biomarkers for tumor recurrence. Furthermore, a subset of advanced MTCs are probably immunogenic, for which single or combined immunotherapy including TIM-3, PD-1, PD-L1, or CTLA-4 blockade may be potential therapeutic approaches in the future.

Near-Infrared Photoimmunotherapy Combined with CTLA4 Checkpoint Blockade in Syngeneic Mouse Cancer Models.

Near infrared photoimmunotherapy (NIR-PIT) is a newly developed and highly selective cancer treatment that induces necrotic/immunogenic cell death. It employs a monoclonal antibody (mAb) conjugated to a photo-absorber dye, IRDye700DX, which is activated by NIR light. Tumor-targeting NIR-PIT is also at least partly mediated by a profound immune response against the tumor. Cytotoxic T-lymphocyte antigen-4 (CTLA4) is widely recognized as a major immune checkpoint protein, which inhibits the immune response against tumors and is therefore, a target for systemic blockade. We investigated the effect of combining tumor-targeted NIR-PIT against the cell-surface antigen, CD44, which is known as a cancer stem cell marker, with a systemic CTLA4 immune checkpoint inhibitor in three syngeneic tumor models (MC38-luc, LL/2, and MOC1). CD44-targeted NIR-PIT combined with CTLA4 blockade showed greater tumor growth inhibition with longer survival compared with CTLA4 blockade alone in all tumor models. NIR-PIT and CTLA4 blockade produced more complete remission in MOC1 tumors (44%) than NIR-PIT and programmed cell death protein 1 (PD-1) blockade (8%), which was reported in our previous paper. However, the combination of NIR-PIT and CTLA4 blockade was less effective in MC38-luc tumors (11%) than the combination of NIR-PIT and PD-1 blockade (70%). Nonetheless, in many cases ineffective results with NIR-PIT and PD-1 blockade were reversed with NIR-PIT and CTLA4 blockade.

SUN-923 Combination Immune Checkpoint Inhibitor Therapy for ACTH-Secreting Pituitary Carcinoma

IntroductionPituitary carcinoma is a rare yet serious entity with poor prognosis despite multimodal therapies. Cerebrospinal and/or systemic metastases are present by definition, making adjuvant systemic therapy necessary. Novel treatments are urgently needed for refractory cases. Immunotherapy with immune checkpoint inhibitors (ICI) targeting cytotoxic T-lymphocyte antigen-4 (CTLA-4), programmed cell death 1 (PD-1) or its ligand (PD-L1) has been a revolution in multiple malignancies. The expression of CTLA-4 and PD-L1 has been elucidated in pituitary adenomas and could be implicated in pituitary carcinomas as well. Hypophysitis is also a frequent endocrine immune-related adverse event, especially during CTLA-4 blockade (with ipilimumab) or combination ICI. However, the efficacy of ICI in the treatment of refractory pituitary tumors has yet to be established. In 2018, Lin et al. successfully treated a first case of a hypermutated aggressive ACTH-secreting pituitary carcinoma with ipilimumab (anti-CTLA-4) and nivolumab (anti-PD-1) combination immunotherapy.Clinical CaseWe report a 40-year old male, diagnosed with an invasive ACTH-secreting pituitary macroadenoma in 2012, initially treated by transsphenoidal and transcranial surgery, followed by adjuvant stereotactic radiotherapy and several courses of ketoconazole. In 2017, he presented to our clinic for a recurrent Cushing’s phenotype despite maximal dosing of ketoconazole. Therapy both with pasireotide and cabergoline was unable to normalize cortisol levels and a bilateral (subtotal) adrenalectomy was performed. In June 2018, he presented to our emergency department with acute diplopia due to a left abducens nerve palsy. Imaging revealed recurrent invasion of the tumor into the sella turcica and cavernous sinus, together with cerebellar and drop metastases at the cervical spine. Temozolomide (TMZ) was initiated for a total of 9 cycles. Progressive disease was observed with development of new onset right oculomotor nerve palsy after the last TMZ cycle, and persistence of elevated serum ACTH-cortisol and urinary cortisol levels, despite the absence of radiological progression. Therefore, he was started in a compassionate use setting with a combination ICI therapy with ipilimumab 3 mg/kg and nivolumab 1 mg/kg (for 4 cycles), followed by maintenance nivolumab therapy (240 mg) every two weeks. He has stable disease (both radiographically and hormonally) five months after the initiation of the immunotherapy.Clinical Lesson(s) or Conclusion(s)We report the second case of ACTH-secreting pituitary carcinoma treated with combination ICI therapy. The disease status of the patient is stable up until now, suggesting at least disease control by the immunotherapy. Checkpoint blockade inhibitors are a promising novel treatment modality for refractory pituitary tumors and should be further studied.

Successful treatment with intralesional talimogene laherparepvec in two patients with immune checkpoint inhibitor-refractory, advanced-stage melanoma

Monoclonal antibodies that block the programmed death-1 (anti-PD-1) or cytotoxic T-lymphocyte antigen-4 (CTLA-4) immune checkpoint receptors (pembrolizumab, nivolumab, ipilimumab, or the combination of nivolumab with ipilimumab) are approved treatment option for patients with advanced melanoma. Over half of all patients are refractory to these immunotherapies and are in need of alternative or complementary treatment options. Talimogene laherparepvec (T-VEC) is a first-in-class intralesionally delivered oncolytic immunotherapy, which has proven efficacy in the treatment of advanced melanoma. A proportion of patients treated with T-VEC will benefit from an abscopal response of noninjected metastases indicative of a systemic antitumor immune response elicited by the intratumoral injections. At present it remains unknown whether the systemic antitumor responses elicited by T-VEC are nonredundant with immune-checkpoint blockade. Recent data on potential synergy between T-VEC and both PD-1 and CTLA-4 blockade suggest that the mechanism of action may be complementary. We report on the successful treatment with intralesional T-VEC of two female patients with locoregionally advanced BRAF V600 wild-type melanoma who previously progressed on anti-PD-1 and anti-CTLA-4 inhibitors.

Effectiveness and safety of PD-1/PD-L1 or CTLA4 inhibitors combined with chemotherapy as a first-line treatment for lung cancer: A meta-analysis.

Immune checkpoint inhibitors (ICIs) combined with chemotherapy have been applied as a first-line treatment for lung cancer, but consistent beneficial results have not been documented. Therefore, our meta-analysis aimed to evaluate the effectiveness and safety of combination therapy to promote its application. We searched electronic databases for studies that estimated the safety and efficacy of combined therapy. The objective response rate (ORR) and disease control response (DCR) parameters were evaluated with odds ratio (OR) values of the combination arm over the non-combination arm. Hazard ratios (HR) and its 95% confidence intervals (95% CI) were used to calculate progression-free survival (PFS) and overall survival (OS) in the combination and non-combination arms. All treatment-related adverse events (TRAEs) and 3 to 5 TRAEs were expressed as relative risk (RR) values of the combination arm over the non-combination arm. Ten eligible studies involving 4,887 patients were identified. The pooled ORs for ORR and DCR were 1.85 (95% CI: 1.30-2.63, P<0.01) and 1.14 (95% CI: 0.70-1.86, P<0.01), respectively. The pooled HRs for PFS and OS were 0.67 (95% CI: 0.58-0.79, P<0.001) and 0.76 (95% CI: 0.65-0.88, P<0.001), respectively. In subgroup analysis, ORR and DCR were significantly improved in the programmed cell death-1/L1 (PD-1/L1) blockade for non-small cell lung cancer (NSCLC) group (subgroup A), with a combined OR values of 2.36 (95% CI: 1.79-3.13, P<0.001) and 1.92 (95% CI: 1.10-3.35, P<0.001), respectively. However, no significant benefits were observed in the cytotoxic T lymphocyte antigen-4 (CTLA-4) blockade for small cell lung cancer (SCLC) (subgroup B) and CTLA-4 blockade for NSCLC groups (subgroup C). In addition, a significant improvement in PFS was observed in subgroup A, subgroup B and subgroup C, with pooled HR values of 0.58 (95% CI: 0.52-0.63, P<0.001), 0.86 (95% CI: 0.76-0.97, P<0.05) and 0.83 (95% CI: 0.68-1.00, P<0.05), respectively. Only subgroup A exhibited an OS benefit, with a combined HR value of 0.67 (95% CI: 0.55-0.81, P<0.001). Moreover, as the expression of PD-L1 increased, the PFS and OS benefits were more significantly. Furthermore, patients without central nervous system (CNS) metastasis who were treated with PD-1/L1 inhibitors had a longer OS than patients with CNS metastasis (HR: 0.67, 95% CI: 0.55-0.80, P<0.001). Finally, combined therapy was associated with 3 to 5 TRAEs (RR: 1.26, 95% CI: 1.08-1.47; P<0.01). Patients treated with immunotherapy and chemotherapy in combination exhibited superior in ORR, DCR, PFS and OS as well as slightly increased TRAE levels compared with those of patients treated with either monotherapy.

Leukemic cell expressed CTLA-4 suppresses T cells via down-modulation of CD80 by trans-endocytosis

Abstract Despite clinical utility in targeting the immune checkpoint, Cytotoxic T Lymphocyte Antigen 4 (CTLA-4), on T cells, its function on non-T cells remains unaddressed, especially in the context of therapy. We define an immunosuppressive role for tumor expressed CTLA-4 in Chronic Lymphocytic Leukemia (CLL). Most primary CLL samples were intracellularly CTLA-4+ (12/14) but surface CTLA-4− (14/14). Co-culture with activated T cells induced surface expression of CTLA-4 on CLL B cells. To mechanistically study CLL B cell expressed CTLA-4, we generated CLL-derived Mec1 and OSU-CLL cell lines to inducibly express CTLA-4. These cell lines express the ligands for CTLA-4, CD80 and CD86. Expression of CTLA-4 on Mec1 and OSU-CLL resulted in decreased CD80 expression on CD80+ cells with rescue upon CTLA-4 blockade (N=3). Co-culture of CTLA-4+ Mec1 and CTLA-4+ primary CLL cells with CD80-GFP+ Hek293 cells revealed transfer of CD80-GFP into primary CLL cells (N=2) and the Mec1 cell line. This finding was consistent with the ability of CTLA-4+ T cells to trans-endocytose CD80. Co-culture of T cells with Mec1 CTLA-4+ cells resulted in decreased IL2 production (N=3, p=0.0172). The loss of IL2 signified decreased co-stimulation as a result of tumor expressed CTLA-4. We generated an in vivo model to study tumor CTLA-4 using the TCL1 murine model of CLL. Murine CTLA-4+ (mCTLA-4) TCL1 tumor cells were engrafted into human CTLA-4+/+mFcγR−/− immune competent mice. MCTLA-4 on the tumor cells but not human CTLA-4 on the host cells was blocked with an anti-mCTLA-4 antibody. Effect on disease progression and overall survival are currently being assessed. We present a paradigm shift whereby CTLA-4 is an immunosuppressive protein irrespective of expression on T cells.

Recent Progress in Mutation-driven Therapy, Immunotherapy and Combination Therapy for the Treatment of Melanoma.

With increases in our understanding of the human genome and immune system, the treatment armamentarium for melanoma has benefitted from the development and approval of BRAF inhibitors, MEK inhibitors, immune checkpoint modulators via cytotoxic T-lymphocyte antigen-4 blockade, and PD-1 and PD-L1 inhibitors. These advances, however, have raised questions about combination therapy, the optimal sequential use of these agents, the limited assessment of response using traditional metrics, and the optimal selection of the population to be treated. In this review we summarize recent breakthroughs and then itemize the development of newer agents, potential prognostic and predictive biomarkers, resistance mechanisms, and strategies of combination therapy. We also emphasize the multifaceted attributes of immunotherapy in terms of durable responses and longterm survival that paradoxically necessitate further research into the underlying mechanisms and longer patient follow-up.

A Phase 1 Study of Nivolumab in Combination with Ipilimumab for Relapsed or Refractory Hematologic Malignancies (CheckMate 039)

Introduction: Nivolumab (nivo) is a fully human IgG4 monoclonal antibody (mAb) targeting programmed death receptor-1 (PD-1). Nivo has demonstrated clinical activity and an acceptable safety profile in a phase 1b study (NCT01592370; CheckMate 039) in patients (pts) with relapsed/refractory hematologic malignancies. In pts diagnosed with Hodgkin lymphoma (HL), after a median 86 weeks of follow-up, 7/20 responders maintained a response for >1.5 years (Ansell S et al. Blood 2015;126:583), and after a median follow-up of 67 weeks, clinical activity (investigator-assessed objective response rate) was demonstrated in follicular lymphoma (FL; 40%), diffuse large B-cell lymphoma (DLBCL; 36%), mycosis fungoides (15%), and peripheral T-cell lymphoma (PTCL; 40%) (Lesokhin AM et al. J Clin Oncol 2016;34:2698). CheckMate 039 also included a cohort of pts who had received nivo in combination with ipilimumab (ipi), a fully human mAb targeting cytotoxic T-lymphocyte antigen 4 (CTLA-4). Combination of CTLA-4 and PD-1 blockade has shown superior efficacy compared with nivo or ipi alone in preclinical studies and solid tumor malignancies (Wolchok JD et al. N Engl J Med 2013;369:122; Larkin JM et al. N Engl J Med 2015;373:22; Antonia SJ et al. Lancet Oncol 2016;17:883). The aim of this cohort study was to evaluate the safety and efficacy of combined immune checkpoint blockade (nivo+ipi) in pts with the following hematologic malignancies: HL, B-cell non-Hodgkin lymphoma (B-NHL; FL and DLBCL), T-cell NHL (T-NHL; cutaneous T-cell lymphoma [CTCL] and PTCL]), and multiple myeloma (MM).Methods: Nivo+ipi were given at 3 mg/kg IV and 1 mg/kg IV, respectively, every 3 weeks for 4 doses, followed by nivo monotherapy (3 mg/kg) every 2 weeks for up to 2 years. Pts with any of the above histologies, relapsed or refractory disease after ≥2 prior lines of therapy, and adequate organ function were included in the study. Prior systemic therapy may have included chemotherapy and autologous hematopoietic stem cell transplantation (auto-HSCT). Prior anti-PD-1 therapy and allogeneic (allo)-HSCT were not permitted. The primary endpoint was safety. Secondary endpoints included investigator-assessed objective response rate (ORR), best overall response, duration of response (DOR), and progression-free survival (PFS).Results: In total, 65 pts were treated with nivo+ipi (31 HL, 15 B-NHL, 11 T-NHL, 7 MM, and 1 with primary mediastinal B-cell lymphoma [PMBL] who was included in the overall safety cohort only). Median (range) number of prior systemic therapies was 4 (2, 10; HL), 3, (1, 16; B-NHL), 4 (1, 11; T-NHL), and 5 (2, 20; MM). Among patients with HL, only 13% (4/31) had prior auto-HSCT. 2 pts with HL and 1 with T-NHL proceeded to allo-HSCT after stopping study therapy. Across all cohorts, median follow-up was 11.4 months. 5 pts (8%) discontinued due to a drug-related adverse event (AE). The most common drug-related AEs of any grade were fatigue (17 pts [26%]), pyrexia (15 [23%]), and diarrhea (12 [18%]). 19 pts (29%) had a drug-related AE of grade ≥3. 31 pts (48%) had a serious AE. 24 pts (37%) died: HL 2 pts, B-NHL 11, T-NHL 6, MM 4, PMBL 1. Among those pts, 22 (34%) were from disease progression (HL 2 pts, B-NHL 10, T-NHL 5, MM 4, PMBL 1); no deaths were due to an AE. Clinical outcome data are presented (Table).Conclusions: These are the first reported data of combination checkpoint blockade therapy in hematologic malignancies. Overall, the combination of nivo+ipi in these heavily pretreated patients demonstrated a safety and efficacy profile similar to that previously reported for nivo monotherapy in HL, NHL, and MM. Additional follow-up may further clarify the role of ipi in this cohort of patients. In this predominantly transplant-naïve group of patients with HL, the efficacy of nivo+ipi was similar to that seen in patients with relapsed/refractory HL treated with nivo alone.Funding: Bristol-Myers Squibb (BMS). Medical writing: S Addison, Caudex, funded by BMS [Display omitted] DisclosuresAnsell:BMS, Seattle Genetics, Merck, Celldex and Affimed: Research Funding. Gutierrez:Bayer Health Care Pharmaceuticals, Inc.: Other: Traveling and Lodging- Food and Beverage; E.R. Squibb & Sons, LLC (Bristol Myers Squibb): Consultancy, Other: Travel and Lodging; Incyte Corporation: Consultancy; Pfizer Inc: Consultancy; Merck Sharp & Dohme Corporation: Consultancy, Other: Travel and Lodging; Pharmacyclics LLC, An AbbVie Company: Other: Food and Beverage. Shipp:Cell Signaling: Honoraria; Bristol-Myers Squibb: Consultancy, Research Funding; Bayer: Research Funding; Merck, Gilead, Takeda: Other: Scientific Advisory Board. Moskowitz:Seattle Genetics: Research Funding; Seattle Genetics, Merck: Consultancy. Borello:Bristol-Myers Squibb: Research Funding, Speakers Bureau. Popa-Mckiver:Bristol-Myers Squibb: Employment, Equity Ownership. Farsaci:Bristol-Myers Squibb: Employment. Zhu:Bristol-Myers Squibb: Employment. Armand:Sequenta Inc: Research Funding; Pfizer: Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Infinity Pharmaceuticals: Consultancy; Roche: Research Funding; Merck: Consultancy, Research Funding.

Ipilimumab and Bevacizumab in Glioblastoma

The median survival in glioblastoma is just over a year, with no standard second-line therapy. Ipilimumab is an immune checkpoint inhibitor that activates the anti-tumour immune response by cytotoxic T-lymphocyte antigen-4 blockade. There is significant evidence supporting its role in the treatment of malignant melanoma, including in patients with brain metastases. The addition of the anti-angiogenesis agent, bevacizumab, seems to offer additional benefit and limit the immune-related side-effects of ipilimumab in melanoma. To date there have been no clinical trials investigating this combination in glioblastoma. In this single practice case series, 20 patients with glioblastoma were consented for and treated with ipilimumab and bevacizumab in combination. Safety, tolerability and the response to treatment were reviewed for all patients. Three patients were treated after palliative first-line radiotherapy, one patient after first-line chemoradiation and 16 patients were treated with recurrent disease. Sixty-five per cent of patients completed four cycles of 3 weekly ipilimumab therapy, administered with 2 weekly bevacizumab. Radiographic responses for patients with recurrent disease were evaluated by Response Assessment in Neuro-oncology (RANO) criteria; 31% of patients showed a partial response, 31% had stable disease and 38% had disease progression. The treatment combination was well tolerated, with treatment terminated before completion due to adverse events in two patients. Autoimmune toxicity was manageable with systemic corticosteroid therapy. Ipilimumab and bevacizumab in combination show promising activity with a predictable and manageable toxicity profile, warranting further clinical studies.

Increased CD4+ T cell proliferation upon treatment correlates with improved overall survival of patients with metastatic melanoma upon anti-CTLA-4 therapy

Abstract Blockade of cytotoxic T-lymphocyte antigen 4 (CTLA-4) augments anti-tumor immunity and has shown success in treating patients with metastatic melanoma. Mechanisms of anti-CTLA-4 efficacy have been proposed in animal models, yet the mechanisms in humans remain to be elucidated. Increased frequency of ICOS+ CD4+ T cells has been shown to be a marker of drug bioactivity, but discovery of prognostic indicators and baseline indicators of which patients are most likely to benefit from CTLA-4 blockade remains a priority. PBMCs from 29 metastatic melanoma patients in a multicenter compassionate use trial of anti-CTLA-4 antibody were evaluated using flow cytometry. At baseline and 12 weeks following initiation of treatment, CTLA-4, HLA-DR, ICOS, PD-L1, PD-1, and Ki-67 frequencies were assessed on CD4+ and CD8+ T cells, as well as CD3+CD4+ regulatory T cells. Independent of survival, anti-CTLA-4 treatment significantly increased the frequency of ICOS+ CD4+ T cells, corroborating previous findings. Interestingly, this increase was also observed in ICOS+ Tregs, but not in CD8+ T cells. Increased proliferative Ki67+ CD4+ T cells and Ki67+ Tregs upon CTLA-4 blockade were both associated with prolonged overall survival. Additionally, when the frequency of Ki67+ CD4+ T cells post-treatment was compared to the baseline frequency, patients with a ratio greater than 1.5 had greater overall survival. Increased post-treatment absolute lymphocyte count—a known prognostic biomarker—also correlated with clinical outcome. Our results suggest that anti-CTLA-4 therapy may be mediated by proliferation of CD4+ T cells, and we identify this proliferation as a novel prognostic biomarker in metastatic melanoma patients treated with anti-CTLA-4 antibody.

The oncolytic peptide LTX-315 overcomes resistance of cancers to immunotherapy with CTLA4 checkpoint blockade.

Intratumoral immunotherapies aim at reducing local immunosuppression, as well as reinstating and enhancing systemic anticancer T-cell functions, without inducing side effects. LTX-315 is a first-in-class oncolytic peptide-based local immunotherapy that meets these criteria by inducing a type of malignant cell death that elicits anticancer immune responses. Here, we show that LTX-315 rapidly reprograms the tumor microenvironment by decreasing the local abundance of immunosuppressive Tregs and myeloid-derived suppressor cells and by increasing the frequency of polyfunctional T helper type 1/type 1 cytotoxic T cells with a concomitant increase in cytotoxic T-lymphocyte antigen-4 (CTLA4) and drop in PD-1 expression levels. Logically, in tumors that were resistant to intratumoral or systemic CTLA4 blockade, subsequent local inoculation of LTX-315 cured the animals or caused tumor regressions with abscopal effects. This synergistic interaction between CTLA4 blockade and LTX-315 was reduced upon blockade of the β-chain of the interleukin-2 receptor (CD122). This preclinical study provides a strong rationale for administering the oncolytic peptide LTX-315 to patients who are receiving treatment with the CTLA4 blocking antibody ipilimumab.

Cytotoxic T lymphocyte antigen-4 expression in esophageal carcinoma: implications for prognosis

To examine the relationship between cytotoxic T lymphocyte antigen-4 (CTLA-4) expression and esophageal carcinoma prognosis, CTLA-4 expression was immunohistochemically detected in paraffin-embedded primary tumor specimens from 158 patients with esophageal cancer. CTLA-4 was detected in the cytoplasm and cell membranes of esophageal cancer cells and in interstitial lymphocytes. In univariate analyses (log-rank), higher interstitial CTLA-4+ lymphocyte density and higher tumor CTLA-4 expression were associated with shorter overall survival (OS). After controlling for age and clinical stage, multivariate analysis (Cox) found that tumor CTLA-4 expression was an independent predictor of shorter OS (HR 2.016, P = 0.004). These results indicate that CTLA-4 expression in the tumor environment (both lymphocytes and tumor cells) is associated with poorer prognosis. In addition, CTLA-4 profiles may be useful for predicting the benefits and toxicity of CTLA-4 blockade in patients with esophageal carcinoma.

Glioblastoma Eradication Following Immune Checkpoint Blockade in an Orthotopic, Immunocompetent Model.

Inhibition of immune checkpoints, including cytotoxic T-lymphocyte antigen-4 (CTLA-4), programmed death-1 (PD-1), and its ligand PD-L1, has demonstrated exciting and durable remissions across a spectrum of malignancies. Combinatorial regimens blocking complementary immune checkpoints further enhance the therapeutic benefit. The activity of these agents for patients with glioblastoma, a generally lethal primary brain tumor associated with significant systemic and microenvironmental immunosuppression, is not known. We therefore systematically evaluated the antitumor efficacy of murine antibodies targeting a broad panel of immune checkpoint molecules, including CTLA-4, PD-1, PD-L1, and PD-L2 when administered as single-agent therapy and in combinatorial regimens against an orthotopic, immunocompetent murine glioblastoma model. In these experiments, we observed long-term tumor-free survival following single-agent anti-PD-1, anti-PD-L1, or anti-CTLA-4 therapy in 50%, 20%, and 15% of treated animals, respectively. Combination therapy of anti-CTLA-4 plus anti-PD-1 cured 75% of the animals, even against advanced, later-stage tumors. In long-term survivors, tumor growth was not seen upon intracranial tumor rechallenge, suggesting that tumor-specific immune memory responses were generated. Inhibitory immune checkpoint blockade quantitatively increased activated CD8(+) and natural killer cells and decreased suppressive immune cells in the tumor microenvironment and draining cervical lymph nodes. Our results support prioritizing the clinical evaluation of PD-1, PD-L1, and CTLA-4 single-agent targeted therapy as well as combination therapy of CTLA-4 plus PD-1 blockade for patients with glioblastoma.

Late administration of murine CTLA-4 blockade prolongs CD8-mediated anti-tumor effects following stimulatory cancer immunotherapy.

We have demonstrated that immunostimulatory therapies such as interleukin-2 (IL-2) and anti-CD40 (αCD40) can be combined to deliver synergistic anti-tumor effects. While this strategy has shown success, efficacy varies depending on a number of factors including tumor type and severe toxicities can be seen. We sought to determine whether blockade of negative regulators such as cytotoxic T lymphocyte antigen-4 (CTLA-4) could simultaneously prolong CD8(+) T cell responses and augment T cell anti-tumor effects. We devised a regimen in which anti-CTLA-4 was administered late so as to delay contraction and minimize toxicities. This late administration both enhanced and prolonged CD8 T cell activation without the need for additional IL-2. The quality of the T cell response was improved with increased frequency of effector/effector memory phenotype cells along with improved lytic ability and bystander expansion. This enhanced CD8 response translated to improved anti-tumor responses both at the primary and metastatic sites. Importantly, toxicities were not exacerbated with combination. This study provides a platform for rational design of immunotherapy combinations to maximize anti-tumor immunity while minimizing toxicities.

Cytotoxic T lymphocyte antigen-4 and immune checkpoint blockade.

The relationship between cancer and the immune system is complex and provides unique therapeutic opportunities. Cytotoxic T lymphocyte antigen-4 (CTLA-4) is a regulatory molecule that suppresses T cell effector function following initial activation by costimulatory signals. Fully human monoclonal antibodies targeting CTLA-4 have been shown to increase T cell function and antitumor responses in patients with advanced metastatic melanoma. Responses observed with such immune checkpoint therapy can follow a different pattern from that seen with cytotoxic chemotherapy or targeted therapy and may continue after therapy is discontinued. In addition, the toxicities that are associated with anti-CTLA-4 therapy may differ from those of conventional therapies and consist of inflammatory events in parts of the body that do not contain cancerous cells. Early recognition of these inflammatory events and intervention is important, and the identification of predictive biomarkers continues to be an unfulfilled need in the field of immunotherapy. Combinatorial approaches with targeted therapies, radiation therapy, chemotherapy, or other immune checkpoint agonists/antagonists have the potential to increase the efficacy of CTLA-4 blockade.