Concise Review: Modulating Cancer Immunity with Hematopoietic Stem and Progenitor Cells.

Hematopoietic stem and progenitor cells (HSPCs) are the progenitor cells that can regenerate the entire blood compartment, including the immune system. Recent studies have unearthed considerable immune-modulating potential of these cells. They can migrate through chemotactic gradients, differentiate into functional immune cells, and crosstalk with immune cells during infections, autoimmune diseases, and cancers. Although the primary role of HSPCs during solid malignancies is considered immunosuppressive, recent studies have discovered immune-activating HSPCs and progeny. In this review, we will discuss the recent evidence that HSPCs act as immunomodulators during solid cancers and highlight the future directions of discovery. Stem Cells 2019;37:166-175.

RNA-Modified T Cells Mediate Effective Delivery of Immunomodulatory Cytokines to Brain Tumors.

With the presence of the blood-brain barrier (BBB), successful immunotherapeutic drug delivery to CNS malignancies remains a challenge. Immunomodulatory agents, such as cytokines, can reprogram the intratumoral microenvironment; however, systemic cytokine delivery has limited access to the CNS. To bypass the limitations of systemically administered cytokines, we investigated if RNA-modified T cells could deliver macromolecules directly to brain tumors. The abilities of T cells to cross the BBB and mediate direct cytotoxic killing of intracranial tumors make them an attractive tool as biological carriers. Using T cell mRNA electroporation, we demonstrated that activated T cells can be modified to secrete granulocyte macrophage colony-stimulating factor (GM-CSF) protein while retaining their inherent effector functions in vitro. GM-CSF RNA-modified T cells effectively delivered GM-CSF to intracranial tumors in vivo and significantly extended overall survival in an orthotopic treatment model. Importantly, GM-CSF RNA-modified T cells demonstrated superior anti-tumor efficacy as compared to unmodified T cells alone or in combination with systemic administration of recombinant GM-CSF. Anti-tumor effects were associated with increased IFN-γ secretion locally within the tumor microenvironment and systemic antigen-specific T cell expansion. These findings demonstrate that RNA-modified T cells may serve as a versatile platform for the effective delivery of biological agents to CNS tumors.

Blast Phase of Myeloproliferative Neoplasm Resembles Acute Myeloid Leukemia, Myelodysplasia-Related, in Clinical Presentation, Cytogenetic Pattern, and Genomic Profile, and Often Undergoes Reversion to Second Chronic Phase Status After Induction Chemotherapy.

CONTEXT.­: BCR::ABL-negative myeloproliferative neoplasm (MPN) has a prolonged clinical course, and some cases eventually undergo transformation to blast phase; its pathogenesis remains to be elucidated. OBJECTIVE.­: To evaluate the clinicopathologic characteristics of MPN in blast phase. DESIGN.­: The study aimed to retrospectively analyze the clinical and laboratory data of 24 cases. RESULTS.­: Median latency to blast phase was 48 months (range, 7-384 months). Complex karyotypes were seen in 12 of the 24 cases (50%). Overall, 16 cases (66.7%) exhibited high allele burdens of MPN driver mutations along with increased blasts, consistent with linear clonal evolution, whereas the remainder (8; 33.3%) showed loss or partial loss of the driver mutation suggestive of a parallel evolution. Additional mutations were noted in 23 cases (100%), including TP53 mutations in 10 of 24 cases (41.7%). Following chemotherapy, 15 of the 24 patients (62.5%) reverted to a second chronic phase while retaining or regaining MPN driver mutations and losing blast-related mutations, although 9 of the 15 patients (60%) later died of disease progression. Median overall survival was 10 months (CI, 4.6-15.4), with those harboring complex karyotypes demonstrating decreased survival (6 versus 29 months; P = .004). CONCLUSIONS.­: MPN-blast phase resembles acute myeloid leukemia, myelodysplasia-related, in cytogenetic pattern, mutation profile, and clinical outcome. Two patterns of clonal evolution are inferred by dynamic analysis of mutation profiles: linear and parallel evolutions. Although overall survival was dismal, 62.5% of our cases achieved second chronic phase, and they showed better survival than those without second chronic phase.

Chronic Lymphocytic Leukemia With Two B-Cell Populations of Discordant Light Chain Restrictions in Individual Patients: Parallel Development of Biclonal B-Cell Neoplasms or Clonal Evolution With Isotype Switch?

OBJECTIVES: To evaluate clinicopathologic characteristics of biclonal chronic lymphocytic leukemia (CLL). METHODS: Retrospectively analyze clinical data and pathologic features. RESULTS: Ten cases were identified in which flow cytometry demonstrated an abnormal B-cell population with a CLL-like immunophenotype but showed no definitive light chain restriction. All had cytogenetic abnormalities detected, including seven with two CLL-related abnormalities. Four of these showed features suggestive of clonal evolution, all having del(13q) as a "stem-line" abnormality and three showing del(11q) as a "side-line" abnormality. Five (50%) cases demonstrated deleterious NOTCH1 mutations, in contrast to 11.8% in a control group of monoclonal CLL (P < .05). Of the 10 patients, 5 received treatment, with good/partial response in three cases and therapeutic resistance in one case. The median treatment-free survival was estimated at 68 months. CONCLUSIONS: Despite a polytypic pattern of light chain expression, the neoplastic nature of biclonal CLL is suggested by a characteristic CLL phenotype and can be confirmed by cytogenetic and genomic analyses. The two clones with discordant light chain isotypes may share a "stem-line" cytogenetic abnormality, suggesting possible clonal evolution. Biclonal CLL is associated with NOTCH1 mutations, which may occur in a small subclone and gradually evolve in clonal size. Genomic analysis on light chain-sorted and/or chronologically collected samples may provide insight into clonal evolution in CLL.

Endometrioid adenofibroma of ovary: A case report and review of literature.

RATIONALE: Endometrioid adenofibroma is a benign epithelial neoplasm of the ovary, most of which are often unilateral. The symptoms of endometrioid adenofibroma are often nonspecific and misleading. Therefore, a full understanding of the characteristics, diagnosis, and treatment methods of this disease is of great importance. In this study, we report a 34-year-old woman who was found with an unidentified mass on the right ovary during the physical examination 3 years ago with nosymptoms or signs. PATIENT CONCERNS: A 34-year-old Chinese female was found with an unidentified 6 cm mass on the right ovary for 3 years that presented with no symptoms or signs. DIAGNOSIS: Pelvic ultrasound revealed a 6 cm cystic solid mixed mass on the right ovary. Through histological and immunohistochemical examinations, the tumor mass was finally diagnosed as endometrioid adenofibroma of ovary. INTERVENTIONS: To confirm the diagnosis, the ovarian tumor was laparoscopically resected. OUTCOMES: The patient returned to hospital after 3 months with no recurrence or postoperative complications. LESSONS: Endometrioid adenofibroma is a benign epithelial neoplasm of the ovary. Complete surgical resection is required and rare cases can recur. Postsurgical pathologic and immunohistochemical testing can confirm a diagnosis of endometrioid adenofibroma. It is important to understand of the key points of differential diagnosis of the disease due to the different prognosis and clinical treatment.

Atypical thymic carcinoid tumor with ectopic ACTH syndrome in a 33-year-old male patient: A rare case report and literature review.

RATIONALE: Atypical thymic carcinoid tumor is an exceedingly rare thymic neuroendocrine tumor derived from the cells of neuroendocrine system. Misdiagnosis or delayed diagnosis may result in disease progression to advanced stages and eventually leads to a poor prognosis. It is therefore necessary to make a correct diagnosis and provide an adequate treatment. PATIENT CONCERNS: A 33-year-old Chinese male presented with numbness in bilateral lower extremities and general fatigue for a month. Chest computed tomography revealed a superior anterior mediastinal mass. Thymoma was initially considered, given the location of the mass and radiographic presentation. DIAGNOSIS: Microscopic findings showed that the tumor cells are arranged in pseudoepitheliomatous growth or irregular nested growth pattern in a background of fibroconnective tissue, with focal infiltration into adipose tissue. The chrysanthemum-like structure or beam-like structure seen often in typical carcinoid tumor was not identified in this case. The tumor cells are spindled or oval, with focal active mitosis. The immunohistochemical staining showed strong positivity for CD56, CgA and Syn, positivity for CK, ACTH, and TTF-1, negativity for Vimentin, and ki67 labeled proliferation index was up to 10% in focal areas. According to the radiological and pathological findings, the diagnosis of atypical thymic carcinoid was made. INTERVENTIONS: The patient underwent surgical resection of the mass. OUTCOME: No recurrence or metastasis was identified during the follow up. LESSONS: Because of its low incidencen, onspecific clinical symptoms, tissue location, and radiological findings, atypical thymic carcinoid tumor may sometimes be misdiagnosed as thymoma. Attention should be paid to avoid misdiagnosis.

Immunotherapy reverses glioma-driven dysfunction of immune system homeostasis.

BACKGROUND: Glioma-induced immune dysregulation of the hematopoietic system has been described in a limited number of studies. In this study, our group further demonstrates that gliomas interrupt the cellular differentiation programming and outcomes of hematopoietic stem and progenitor cells (HSPCs) in the bone marrow. HSPCs from glioma-bearing mice are reprogrammed and driven towards expansion of myeloid lineage precursors and myeloid-derived suppressor cells (MDSCs) in secondary lymphoid organs. However, we found this expansion is reversed by immunotherapy. Adoptive cellular therapy (ACT) has been demonstrably efficacious in multiple preclinical models of central nervous system (CNS) malignancies, and here we describe how glioma-induced dysfunction is reversed by this immunotherapeutic platform. METHODS: The impact of orthotopic KR158B-luc glioma on HSPCs was evaluated in an unbiased fashion using single cell RNAseq (scRNAseq) of lineage- cells and phenotypically using flow cytometry. Mature myeloid cell frequencies and function were also evaluated using flow cytometry. Finally, ACT containing total body irradiation, tumor RNA-pulsed dendritic cells, tumor-reactive T cells and HSPCs isolated from glioma-bearing or non-tumor-bearing mice were used to evaluate cell fate differentiation and survival. RESULTS: Using scRNAseq, we observed an altered HSPC landscape in glioma-bearing versus non-tumor-bearing mice . In addition, an expansion of myeloid lineage subsets, including granulocyte macrophage precursors (GMPs) and MDSCs, were observed in glioma-bearing mice relative to non-tumor-bearing controls. Furthermore, MDSCs from glioma-bearing mice demonstrated increased suppressive capacity toward tumor-specific T cells as compared with MDSCs from non-tumor-bearing hosts. Interestingly, treatment with ACT overcame these suppressive properties. When HSPCs from glioma-bearing mice were transferred in the context of ACT, we observed significant survival benefit and long-term cures in orthotopic glioma models compared with mice treated with ACT using non-glioma-bearing HSPCs.

Myelopoiesis during Solid Cancers and Strategies for Immunotherapy.

Our understanding of the relationship between the immune system and cancers has undergone significant discovery recently. Immunotherapy with T cell therapies and checkpoint blockade has meaningfully changed the oncology landscape. While remarkable clinical advances in adaptive immunity are occurring, modulation of innate immunity has proven more difficult. The myeloid compartment, including macrophages, neutrophils, and dendritic cells, has a significant impact on the persistence or elimination of tumors. Myeloid cells, specifically in the tumor microenvironment, have direct contact with tumor tissue and coordinate with tumor-reactive T cells to either stimulate or antagonize cancer immunity. However, the myeloid compartment comprises a broad array of cells in various stages of development. In addition, hematopoietic stem and progenitor cells at various stages of myelopoiesis in distant sites undergo significant modulation by tumors. Understanding how tumors exert their influence on myeloid progenitors is critical to making clinically meaningful improvements in these pathways. Therefore, this review will cover recent developments in our understanding of how solid tumors modulate myelopoiesis to promote the formation of pro-tumor immature myeloid cells. Then, it will cover some of the potential avenues for capitalizing on these mechanisms to generate antitumor immunity.

Immune Escape After Adoptive T-cell Therapy for Malignant Gliomas.

PURPOSE: Immunotherapy has been demonstrably effective against multiple cancers, yet tumor escape is common. It remains unclear how brain tumors escape immunotherapy and how to overcome this immune escape. EXPERIMENTAL DESIGN: We studied KR158B-luc glioma-bearing mice during treatment with adoptive cellular therapy (ACT) with polyclonal tumor-specific T cells. We tested the immunogenicity of primary and escaped tumors using T-cell restimulation assays. We used flow cytometry and RNA profiling of whole tumors to further define escape mechanisms. To treat immune-escaped tumors, we generated escape variant-specific T cells through the use of escape variant total tumor RNA and administered these cells as ACT. In addition, programmed cell death protein-1 (PD-1) checkpoint blockade was studied in combination with ACT. RESULTS: Escape mechanisms included a shift in immunogenic tumor antigens, downregulation of MHC class I, and upregulation of checkpoint molecules. Polyclonal T cells specific for escape variants displayed greater recognition of escaped tumors than primary tumors. When administered as ACT, these T cells prolonged median survival of escape variant-bearing mice by 60%. The rational combination of ACT with PD-1 blockade prolonged median survival of escape variant glioma-bearing mice by 110% and was dependent upon natural killer cells and T cells. CONCLUSIONS: These findings suggest that the immune landscape of brain tumors are markedly different postimmunotherapy yet can still be targeted with immunotherapy.

Effect of atorvastatin on humoral immune response to 23-valent pneumococcal polysaccharide vaccination in healthy volunteers: The StatVax randomized clinical trial.

BACKGROUND: The immunomodulatory effects of statins on vaccine response remain uncertain. Therefore, the objective of this study was to determine if atorvastatin enhances pneumococcal-specific antibody titer following 23-valent pneumococcal polysaccharide vaccination. METHODS: Double-blind, placebo-controlled, single-center randomized clinical trial entitled StatVax. Subjects were enrolled between June and July 2014 and followed up through September 2014. 33 healthy volunteers signed informed consent after volunteer sampling. 11 participants were excluded; 22 healthy volunteers without prior pneumococcal vaccination were enrolled and completed the study. Participants were randomized to receive a 28-day course of 40 mg atorvastatin (n = 12) or matching lactose placebo (n = 10). On day 7 of treatment, Pneumovax 23 was administered intramuscularly. The primary outcome was fold change in total pneumococcal-specific antibody titer determined by a ratio of post-vaccination titer over baseline titer. Secondary outcomes included serotype-specific pneumococcal antibody titer, seroconversion, complete blood counts (CBC), erythrocyte sedimentation rate (ESR), and serum cytokine analysis. RESULTS: Of the 22 randomized patients (mean age, 23.86; SD, 4.121; 11 women [50%]), 22 completed the trial. Total anti-pneumococcal antibody titer in the atorvastatin group went from a baseline mean of 32.58 (SD, 15.96) to 147.7 (SD, 71.52) µg/mL at 21 days post-vaccination while titer in the placebo group went from a mean of 30.81 (SD, 13.04) to 104.4 (SD, 45) µg/mL. When comparing fold change between treatment groups, there was a significant increase in fold change of total anti-pneumococcal antibody titer in the atorvastatin group compared to the placebo group (2-way ANOVA, p = .0177). CONCLUSIONS: Atorvastatin enhances antigen-specific primary humoral immune response to a T cell-independent pneumonia vaccination. Pending confirmation by larger cohort studies of target populations, peri-vaccination conventional doses of statins can become a novel adjuvant for poorly-immunogenic polysaccharide-based vaccines. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT02097589.

Dendritic Cell-Activating Magnetic Nanoparticles Enable Early Prediction of Antitumor Response with Magnetic Resonance Imaging.

Cancer vaccines initiate antitumor responses in a subset of patients, but the lack of clinically meaningful biomarkers to predict treatment response limits their development. Here, we design multifunctional RNA-loaded magnetic liposomes to initiate potent antitumor immunity and function as an early biomarker of treatment response. These particles activate dendritic cells (DCs) more effectively than electroporation, leading to superior inhibition of tumor growth in treatment models. Inclusion of iron oxide enhances DC transfection and enables tracking of DC migration with magnetic resonance imaging (MRI). We show that T2*-weighted MRI intensity in lymph nodes is a strong correlation of DC trafficking and is an early predictor of antitumor response. In preclinical tumor models, MRI-predicted "responders" identified 2 days after vaccination had significantly smaller tumors 2-5 weeks after treatment and lived 73% longer than MRI-predicted "nonresponders". These studies therefore provide a simple, scalable nanoparticle formulation to generate robust antitumor immune responses and predict individual treatment outcome with MRI.

Lin-CCR2+ hematopoietic stem and progenitor cells overcome resistance to PD-1 blockade.

Immune checkpoint blockade using anti-PD-1 monoclonal antibodies has shown considerable promise in the treatment of solid tumors, but brain tumors remain notoriously refractory to treatment. In CNS malignancies that are completely resistant to PD-1 blockade, we found that bone marrow-derived, lineage-negative hematopoietic stem and progenitor cells (HSCs) that express C-C chemokine receptor type 2 (CCR2+) reverses treatment resistance and sensitizes mice to curative immunotherapy. HSC transfer with PD-1 blockade increases T-cell frequency and activation within tumors in preclinical models of glioblastoma and medulloblastoma. CCR2+HSCs preferentially migrate to intracranial brain tumors and differentiate into antigen-presenting cells within the tumor microenvironment and cross-present tumor-derived antigens to CD8+ T cells. HSC transfer also rescues tumor resistance to adoptive cellular therapy in medulloblastoma and glioblastoma. Our studies demonstrate a novel role for CCR2+HSCs in overcoming brain tumor resistance to PD-1 checkpoint blockade and adoptive cellular therapy in multiple invasive brain tumor models.

Cross-talk between T Cells and Hematopoietic Stem Cells during Adoptive Cellular Therapy for Malignant Glioma.

Purpose: Adoptive T-cell immunotherapy (ACT) has emerged as a viable therapeutic for peripheral and central nervous system (CNS) tumors. In peripheral cancers, optimal efficacy of ACT is reliant on dendritic cells (DCs) in the tumor microenvironment. However, the CNS is largely devoid of resident migratory DCs to function as antigen-presenting cells during immunotherapy. Herein, we demonstrate that cellular interactions between adoptively transferred tumor-reactive T cells and bone marrow-derived hematopoietic stem and progenitor cells (HSPCs) lead to the generation of potent intratumoral DCs within the CNS compartment.Experimental Design: We evaluated HSPC differentiation during ACT in vivo in glioma-bearing hosts and HSPC proliferation and differentiation in vitro using a T-cell coculture system. We utilized FACS, ELISAs, and gene expression profiling to study the phenotype and function of HSPC-derived cells ex vivo and in vivo To demonstrate the impact of HSPC differentiation and function on antitumor efficacy, we performed survival experiments.Results: Transfer of HSPCs with concomitant ACT led to the production of activated CD86+CD11c+MHCII+ cells consistent with DC phenotype and function within the brain tumor microenvironment. These intratumoral DCs largely supplanted abundant host myeloid-derived suppressor cells. We determined that during ACT, HSPC-derived cells in gliomas rely on T-cell-released IFNγ to differentiate into DCs, activate T cells, and reject intracranial tumors.Conclusions: Our data support the use of HSPCs as a novel cellular therapy. Although DC vaccines induce robust immune responses in the periphery, our data demonstrate that HSPC transfer uniquely generates intratumoral DCs that potentiate T-cell responses and promote glioma rejection in situClin Cancer Res; 24(16); 3955-66. ©2018 AACR.