rWTC-MBTA Vaccine Induces Potent Adaptive Immune Responses Against Glioblastomas via Dynamic Activation of Dendritic Cells.

Despite strides in immunotherapy, glioblastoma multiforme (GBM) remains challenging due to low inherent immunogenicity and suppressive tumor microenvironment. Converting "cold" GBMs to "hot" is crucial for immune activation and improved outcomes. This study comprehensively characterized a therapeutic vaccination strategy for preclinical GBM models. The vaccine consists of Mannan-BAM-anchored irradiated whole tumor cells, Toll-like receptor ligands [lipoteichoic acid (LTA), polyinosinic-polycytidylic acid (Poly (I:C)), and resiquimod (R-848)], and anti-CD40 agonistic antibody (rWTC-MBTA). Intracranial GBM models (GL261, SB28 cells) are used to evaluate the vaccine efficacy. A substantial number of vaccinated mice exhibited complete regression of GBM tumors in a T-cell-dependent manner, with no significant toxicity. Long-term tumor-specific immune memory is confirmed upon tumor rechallenge. In the vaccine-draining lymph nodes of the SB28 model, rWTC-MBTA vaccination triggered a major rise in conventional dendritic cell type 1 (cDC1) 12 h post-treatment, followed by an increase in conventional dendritic cell type 2 (cDC2), monocyte-derived dendritic cell (moDC), and plasmacytoid dendritic cell (pDC) on Day 5 and Day 13. Enhanced cytotoxicity of CD4+ and CD8+ T cells in vaccinated mice is verified in co-culture with tumor cells. Analyses of immunosuppressive signals (T-cell exhaustion, myeloid-derived suppressor cells (MDSC), M2 macrophages) in the GBM microenvironment suggest potential combinations with other immunotherapies for enhanced efficacy. In conclusion, the authors findings demonstrate that rWTC-MBTA induces potent and long-term adaptive immune responses against GBM.

rWTC-MBTA: autologous vaccine prevents metastases via antitumor immune responses.

BACKGROUND: Autologous tumor cell-based vaccines (ATVs) aim to prevent and treat tumor metastasis by activating patient-specific tumor antigens to induce immune memory. However, their clinical efficacy is limited. Mannan-BAM (MB), a pathogen-associated molecular pattern (PAMP), can coordinate an innate immune response that recognizes and eliminates mannan-BAM-labeled tumor cells. TLR agonists and anti-CD40 antibodies (TA) can enhance the immune response by activating antigen-presenting cells (APCs) to present tumor antigens to the adaptive immune system. In this study, we investigated the efficacy and mechanism of action of rWTC-MBTA, an autologous whole tumor cell vaccine consisting of irradiated tumor cells (rWTC) pulsed with mannan-BAM, TLR agonists, and anti-CD40 antibody (MBTA), in preventing tumor metastasis in multiple animal models. METHODS: The efficacy of the rWTC-MBTA vaccine was evaluated in mice using breast (4T1) and melanoma (B16-F10) tumor models via subcutaneous and intravenous injection of tumor cells to induce metastasis. The vaccine's effect was also assessed in a postoperative breast tumor model (4T1) and tested in autologous and allogeneic syngeneic breast tumor models (4T1 and EMT6). Mechanistic investigations included immunohistochemistry, immunophenotyping analysis, ELISA, tumor-specific cytotoxicity testing, and T-cell depletion experiments. Biochemistry testing and histopathology of major tissues in vaccinated mice were also evaluated for potential systemic toxicity of the vaccine. RESULTS: The rWTC-MBTA vaccine effectively prevented metastasis and inhibited tumor growth in breast tumor and melanoma metastatic animal models. It also prevented tumor metastasis and prolonged survival in the postoperative breast tumor animal model. Cross-vaccination experiments revealed that the rWTC-MBTA vaccine prevented autologous tumor growth, but not allogeneic tumor growth. Mechanistic data demonstrated that the vaccine increased the percentage of antigen-presenting cells, induced effector and central memory cells, and enhanced CD4+ and CD8+ T-cell responses. T-cells obtained from mice that were vaccinated displayed tumor-specific cytotoxicity, as shown by enhanced tumor cell killing in co-culture experiments, accompanied by increased levels of Granzyme B, TNF-α, IFN-γ, and CD107a in T-cells. T-cell depletion experiments showed that the vaccine's antitumor efficacy depended on T-cells, especially CD4+ T-cells. Biochemistry testing and histopathology of major tissues in vaccinated mice revealed negligible systemic toxicity of the vaccine. CONCLUSION: The rWTC-MBTA vaccine demonstrated efficacy in multiple animal models through T-cell mediated cytotoxicity and has potential as a therapeutic option for preventing and treating tumor metastasis with minimal systemic toxicity.

Targeting selenoprotein H in the nucleolus suppresses tumors and metastases by Isovalerylspiramycin I.

BACKGROUND: Compared to normal cells, cancer cells exhibit a higher level of oxidative stress, which primes key cellular and metabolic pathways and thereby increases their resilience under oxidative stress. This higher level of oxidative stress also can be exploited to kill tumor cells while leaving normal cells intact. In this study we have found that isovalerylspiramycin I (ISP I), a novel macrolide antibiotic, suppresses cancer cell growth and tumor metastases by targeting the nucleolar protein selenoprotein H (SELH), which plays critical roles in keeping redox homeostasis and genome stability in cancer cells. METHODS: We developed ISP I through genetic recombination and tested the antitumor effects using primary and metastatic cancer models. The drug target was identified using the drug affinity responsive target stability (DARTS) and mass spectrum assays. The effects of ISP I were assessed for reactive oxygen species (ROS) generation, DNA damage, R-loop formation and its impact on the JNK2/TIF-IA/RNA polymerase I (POLI) transcription pathway. RESULTS: ISP I suppresses cancer cell growth and tumor metastases by targeting SELH. Suppression of SELH induces accumulation of ROS and cancer cell-specific genomic instability. The accumulation of ROS in the nucleolus triggers nucleolar stress and blocks ribosomal RNA transcription via the JNK2/TIF-IA/POLI pathway, causing cell cycle arrest and apoptosis in cancer cells. CONCLUSIONS: We demonstrated that ISP I links cancer cell vulnerability to oxidative stress and RNA biogenesis by targeting SELH. This suggests a potential new cancer treatment paradigm, in which the primary therapeutic agent has minimal side-effects and hence may be useful for long-term cancer chemoprevention.

High-Dose Rate Interstitial Spine Brachytherapy Using an Intraoperative Mobile Computed Tomography-Guided Surgical Navigation System.

BACKGROUND: Up to 15% of previously irradiated metastatic spine tumors will progress. Re-irradiation of these tumors poses a significant risk of exceeding the radiation tolerance to the spinal cord. High-dose rate (HDR) brachytherapy is a treatment alternative. OBJECTIVE: To develop a novel HDR spine brachytherapy technique using an intraoperative computed tomography-guided navigation (iCT navigation). METHODS: Patients with progressive metastatic spine tumors were included in the study. HDR brachytherapy catheters were placed under iCT navigation. CT-based planning with magnetic resonance imaging fusion was performed to ensure conformal dose delivery to the target while sparing normal tissue, including the spinal cord. Patients received single fraction radiation treatment. RESULTS: Five patients with thoracolumbar tumors were treated with HDR brachytherapy. Four patients previously received radiotherapy to the same spinal level. Preimplant plans demonstrated median clinical target volume (CTV) D90 of 116.5% (110.8%-147.7%), V100 of 95.7% (95.5%-99.6%), and Dmax of 8.08 Gy (7.65-9.8 Gy) to the spinal cord/cauda equina. Postimplant plans provided median CTV D90 of 113.8% (93.6%-120.1%), V100 of 95.9% (87%-99%), and Dmax of 9.48 Gy (6.5-10.3 Gy) to cord/cauda equina. Patients who presented with back pain (n = 3) noted symptomatic improvement at a median follow-up of 22 d after treatment. Four patients demonstrated local tumor control of spinal metastatic tumor at a median follow-up of 92 d after treatment. One patient demonstrated radiographic evidence of local tumor progression 2.7 mo after treatment. CONCLUSION: HDR spine brachytherapy with iCT navigation is a promising treatment alternative to induce local tumor control and reduce pain symptoms associated with metastatic spine disease.

Identification of Immune Cell Infiltration in Murine Pheochromocytoma during Combined Mannan-BAM, TLR Ligand, and Anti-CD40 Antibody-Based Immunotherapy.

Immunotherapy has become an essential component in cancer treatment. However, the majority of solid metastatic cancers, such as pheochromocytoma, are resistant to this approach. Therefore, understanding immune cell composition in primary and distant metastatic tumors is important for therapeutic intervention and diagnostics. Combined mannan-BAM, TLR ligand, and anti-CD40 antibody-based intratumoral immunotherapy (MBTA therapy) previously resulted in the complete eradication of murine subcutaneous pheochromocytoma and demonstrated a systemic antitumor immune response in a metastatic model. Here, we further evaluated this systemic effect using a bilateral pheochromocytoma model, performing MBTA therapy through injection into the primary tumor and using distant (non-injected) tumors to monitor size changes and detailed immune cell infiltration. MBTA therapy suppressed the growth of not only injected but also distal tumors and prolonged MBTA-treated mice survival. Our flow cytometry analysis showed that MBTA therapy led to increased recruitment of innate and adaptive immune cells in both tumors and the spleen. Moreover, adoptive CD4+ T cell transfer from successfully MBTA-treated mice (i.e., subcutaneous pheochromocytoma) demonstrates the importance of these cells in long-term immunological memory. In summary, this study unravels further details on the systemic effect of MBTA therapy and its use for tumor and metastasis reduction or even elimination.

Mannan-BAM, TLR Ligands, Anti-CD40 Antibody (MBTA) Vaccine Immunotherapy: A Review of Current Evidence and Applications in Glioblastoma.

The foundation of precision immunotherapy in oncology is rooted in computational biology and patient-derived sample sequencing to enrich for and target immunogenic epitopes. Discovery of these tumor-specific epitopes through tumor sequencing has revolutionized patient outcomes in many types of cancers that were previously untreatable. However, these therapeutic successes are far from universal, especially with cancers that carry high intratumoral heterogeneity such as glioblastoma (GBM). Herein, we present the technical aspects of Mannan-BAM, TLR Ligands, Anti-CD40 Antibody (MBTA) vaccine immunotherapy, an investigational therapeutic that potentially circumvents the need for in silico tumor-neoantigen enrichment. We then review the most promising GBM vaccination strategies to contextualize the MBTA vaccine. By reviewing current evidence using translational tumor models supporting MBTA vaccination, we evaluate the underlying principles that validate its clinical applicability. Finally, we showcase the translational potential of MBTA vaccination as a potential immunotherapy in GBM, along with established surgical and immunologic cancer treatment paradigms.

Metformin enhances anti-cancer effects of cisplatin in meningioma through AMPK-mTOR signaling pathways.

Cisplatin is currently used to treat inoperable recurrent meningiomas, but its side effects and drug resistance limit its use. Metformin has recently been identified as a chemosensitizing agent. However, the combined treatment of cisplatin and metformin in high-grade meningiomas has not been reported. Herein, our findings demonstrate metformin significantly enhanced cisplatin-induced inhibition of proliferation in meningioma cells, which was associated with the induction of G0/G1 cell cycle arrest. Additionally, metformin activated adenosine monophosphate activated protein kinase (AMPK) and repressed the mammalian target of rapamycin (mTOR) signaling pathways via an AMPK-dependent mechanism. Furthermore, our xenograft murine model confirmed that metformin enhanced cisplatin's anti-cancer effect by upregulation of AMPK and downregulation of mTOR signaling pathways. In addition, in 63 patients with atypical meningiomas, the activation of AMPK was significantly associated with tumor recurrence and short disease-free survival (DFS). These results demonstrate metformin enhanced the anti-cancer effect of cisplatin in meningioma in vitro and in vivo, an effect mediated through the activation of AMPK and repression of mTOR signaling pathways. Our study suggests the combined treatment of metformin and cisplatin is an effective and safe treatment for high-grade meningiomas.

Inhibition of PP2A with LB-100 Enhances Efficacy of CAR-T Cell Therapy Against Glioblastoma.

Chimeric antigen receptor (CAR)-engineered T cells represent a promising modality for treating glioblastoma. Recently, we demonstrated that CAR-T cells targeting carbonic anhydrase IX (CAIX), a protein involved in HIF-1a hypoxic signaling, is a promising CAR-T cell target in an intracranial murine glioblastoma model. Anti-CAIX CAR-T cell therapy is limited by its suboptimal activation within the tumor microenvironment. LB-100, a small molecular inhibitor of protein phosphatase 2A (PP2A), has been shown to enhance T cell anti-tumor activity through activation of the mTOR signaling pathway. Herein, we investigated if a treatment strategy consisting of a combination of LB-100 and anti-CAIX CAR-T cell therapy produced a synergistic anti-tumor effect. Our studies demonstrate that LB-100 enhanced anti-CAIX CAR-T cell treatment efficacy in vitro and in vivo. Our findings demonstrate the role of LB-100 in augmenting the cytotoxic activity of anti-CAIX CAR-T cells and underscore the synergistic therapeutic potential of applying combination LB-100 and CAR-T Cell therapy to other solid tumors.

Induction of Immune Response Against Metastatic Tumors via Vaccination of Mannan-BAM, TLR Ligands and Anti-CD40 Antibody (MBTA).

Emerging evidence is demonstrating the extent of T-cell infiltration within the tumor microenvironment has favorable prognostic and therapeutic implications. Hence, immunotherapeutic strategies that augment the T-cell signature of tumors hold promising therapeutic potential. Recently, immunotherapy based on intratumoral injection of mannan-BAM, toll-like receptor ligands and anti-CD40 antibody (MBTA) demonstrated promising potential to modulate the immune phenotype of injected tumors. The strategy promotes the phagocytosis of tumor cells to facilitate the recognition of tumor antigens and induce a tumor-specific adaptive immune response. Using a syngeneic colon carcinoma model, we demonstrate MBTA's potential to augment CD8+ T-cell tumor infiltrate when administered intratumorally or subcutaneously as part of a whole tumor cell vaccine. Both immunotherapeutic strategies proved effective at controlling tumor growth, prolonged survival and induced immunological memory against the parental cell line. Collectively, our investigation demonstrates MBTA's potential to trigger a potent anti-tumor immune response.

Comparison of Clinical Outcomes Stratified by Target Delineation for Patients Undergoing Stereotactic Body Radiotherapy for Spinal Metastases.

OBJECTIVE: Stereotactic body radiotherapy (SBRT) is an effective treatment of spinal metastases in the vertebral body. However, variation has existed between practitioners regarding the appropriate target delineation. As such, we compared the tumor control, rates of compression fractures, and pain control for patients who had undergone SBRT for spinal metastases to either the lesion only (LO) or the full vertebral body (FVB). METHODS: A total of 126 spinal metastases in 84 patients had received single-fraction SBRT from January 2009 to February 2015. Of the 126 lesions, 36 (29%) were in the FVB group and 90 were in the LO group. The SBRT plans were reviewed to determine the treatment volume. Odds ratios were used to compare the rates of compression fracture and local failure. Regression analysis was performed to identify the predictors of outcome. RESULTS: A total of 5 failures had occurred in the FVB group and 14 in the LO group; however, the difference was not statistically significant (P = 0.5). No difference was found in pain reduction between the 2 groups (P = 0.9). Seven post-treatment compression fractures occurred in the LO group and four in the FVB group; however, the difference was not statistically significant (P = 0.6). The minimum dose to the planning target volume, patient age, and planning target volume size were the only significant factors predicting for local failure, vertebral body fracture, and pain control, respectively. CONCLUSIONS: Given that we found no difference in tumor control, pain reduction, or fracture rate between patients treated to the FVB versus the. LO, it might be reasonable to consider SBRT to the LO for select patients.

Blocking lncRNA MALAT1/miR-199a/ZHX1 Axis Inhibits Glioblastoma Proliferation and Progression.

Zinc fingers and homeoboxes 1 (ZHX1) is a transcription repressor that has been implicated in the tumorigenesis and progression of diverse tumors. The functional role and regulating mechanism of ZHX1 has not been elucidated in glioblastoma (GBM). Previous reports have suggested that a large number of non-coding RNAs play a vital role in glioma initiation and progression. This study aimed to investigate the functional role and co-regulatory mechanisms of the metastasis-associated lung adenocarcinoma transcript-1 (MALAT1)/ microRNA-199a (miR-199a)/ZHX1 axis in GBM. We analyzed the expression of the MALAT1/miR-199a/ZHX1 axis and its correlation with patients' overall survival using two different glioma gene-expression datasets. A series of in vitro and in vivo studies including dual luciferase reporter assay, fluorescence in situ hybridization (FISH), RNA immunoprecipitation, and pull-down experiments were completed to elucidate the biological significance of the MALAT1/miR-199a/ZHX1 axis in promoting glioma proliferation and progression. Elevated ZHX1 expression correlated with poor prognosis in GBM patients, and in vitro studies demonstrated that ZHX1 attenuated GBM cell apoptosis by downregulation of pro-apoptotic protein (Bax) and upregulation of anti-apoptotic protein (Bcl-2). Furthermore, knockdown of MALAT1 inhibited GBM proliferation and progression in vitro and reduced tumor volume and prolonged survival in an orthotopic GBM murine model. Finally, we demonstrated that MALAT1 promoted ZHX1 expression via acting as a competing endogenous RNA by sponging miR-199a. The MALAT1/miR-199a/ZHX1 axis promotes GBM cell proliferation and progression in vitro and in vivo, and its expression negatively correlates with GBM patient survival. Blocking the MALAT1/miR-199a/ZHX1 axis can serve as a novel therapeutic strategy for treating GBM.

Natural history of the aging spine: a cross-sectional analysis of spinopelvic parameters in the asymptomatic population.

OBJECTIVE: There have been numerous studies demonstrating increased pain and disability when patients' spinopelvic parameters fall outside of certain accepted ranges. However, these values were established based on patients suffering from spinal deformities. It remains unknown how these parameters change over a lifetime in asymptomatic individuals. The goal of this study was to define a range of spinopelvic parameters from asymptomatic individuals. METHODS: Sagittal scoliosis radiographs of 210 asymptomatic patients were evaluated. All measurements were reviewed by 2 trained observers, supervised by a trained clinician. The following parameters and relationships were measured or calculated: cervical lordosis (CL), thoracic kyphosis (TK), lumbar lordosis (LL), pelvic incidence (PI), sagittal vertical axis (SVA), cervical SVA (cSVA), and T1 slope, TK/LL, truncal inclination, pelvic tilt (PT), LL-PI, LL/PI, and T1 slope/PI. Patients were stratified by decade of life, and regression analysis was performed to delineate the relationship between each consecutive age group and the aforementioned parameters. RESULTS: Cervical lordosis (R2 = 0.61), thoracic kyphosis (R2 = 0.84), SVA (R2 = 0.88), cSVA (R2 = 0.51), and T1 slope (R2 = 0.77) all increase with age. Truncal inclination (R2 = 0.36) and T1 slope/CL remain stable over all decades (R2 = 0.01). LL starts greater than PI, but in the 6th decade of life, LL becomes equal to PI and in the 7th decade becomes smaller than PI (R2 = 0.96). The ratio of TK/LL is stable until the 7th decade of life (R2 = 0.81), whereas PT is stable until the 6th decade (R2 = 0.92). CONCLUSIONS: This study further refines the generally accepted LL = PI + 10° by showing that patients under the age of 50 years should have more LL compared to PI, whereas after the 5th decade the relationship is reversed. SVA was not as sensitive across age groups, exhibiting a marked increase only in the 7th decade of life. Given the reliable increase of CL with age, and the stability of T1 slope/CL, this represents another important relationship that should be maintained when performing cervical deformity/fusion surgery. This study has important implications for evaluating adult patients with spinal deformities and for establishing corrective surgical goals.

Blocking lncRNA H19-miR-19a-Id2 axis attenuates hypoxia/ischemia induced neuronal injury.

Elevated expression of lncRNA H19 (H19) in the setting of hypoxia has been implicated as a promising therapeutic target for various cancers. However, little is known about the impact and underlying mechanism of H19 in ischemic brain stroke. This study found that H19 levels were elevated in the serum of stroke patients, as well as in the ischemic penumbra of rats with middle cerebral artery occlusion/reperfusion (MCAO/R) injury and neuronal cells with oxygen glucose deprivation (OGD). Further, knockdown of H19 with siRNA alleviated cell apoptosis in OGD neuronal cells, and inhibition of H19 in MCAO/R rats significantly decreased neurological deficit, brain infarct volume and neuronal apoptosis. Lastly, with gain and loss of function studies, dual luciferase reported assay, RNA immunoprecipitation (RIP) and pull-down experiments, we demonstrated the dual competitive interaction of miR-19a with H19 and the 3'-UTR of Id2 mRNA, resulting in the identification of the H19-miR-19a-Id2 axis. With biological studies, we also revealed that H19-miR-19a-Id2 axis modulated hypoxia induced neuronal apoptosis. This study demonstrates that the identified H19-miR-19a-Id2 axis plays a critical role in hypoxia induced neuronal apoptosis, and blocking this axis may serve as a novel therapeutic strategy for ischemic brain injury.

Development of a sickle cell disease readiness for transition assessment.

BACKGROUND: Instrumentation with established reliability and validity is not yet routinely utilized to assess readiness for transition from pediatric to adult care for youth and young adults with chronic conditions, including sickle cell disease (SCD). OBJECTIVE: The aim of this study was to develop a SCD specific readiness for transition assessment tool. SUBJECTS: Fifty-seven youths with SCD, ages 15-21 years, completed the initial version of the Transition Intervention Program - Readiness for Transition (TIP-RFT) assessment; 113 youths/young adults with SCD, ages 14-26 years, at two distinct sites of care completed a refined version of the TIP-RFT. METHODS: The TIP-RFT was constructed based on a literature review, provider and patient consensus and assessed domains including knowledge and skills in medical self-care, social support, health benefits and independent living and educational/vocational skills. We used principal components factor analysis to evaluate TIP-RFT responses and assessed differences in TIP-RFT scores in relation to age, gender, sickle cell diagnosis and site of care. RESULTS: The original TIP-RFT, which had demonstrated face validity, was reduced from 56 to 22 items. The revised instrument consisting of four subscales demonstrated good internal consistency reliability and construct validity. CONCLUSION: Our results support that the TIP-RFT is a valid and reliable tool for the assessment of transition readiness for youths with SCD. The TIP-RFT assessment can guide interventions to improve transition readiness and can provide a foundation for future research on other variables that might be associated with transition readiness.

Self-efficacy and readiness for transition from pediatric to adult care in sickle cell disease.

BACKGROUND: Theories of self-care management, particularly the development of self-efficacy or confidence in one's ability to manage health-related goals, tasks, and challenges may provide a useful framework for developing programs to improve transition from pediatric to adult care for youth and young adults with sickle cell disease (SCD). OBJECTIVE: The aim of this study was to evaluate the hypothesis stating that ratings of self-efficacy is positively associated with self-ratings of transition readiness. SUBJECTS: A total of 113 individuals with SCD aged 14-26 years at two distinct sites of care were recruited for the study. MATERIALS AND METHODS: Participants completed the Transition Intervention Program Readiness for Transition (TIP-RFT) assessment, the Sickle Cell Self-Efficacy Scale and the Sickle Cell Stress -Adolescent scale. RESULTS: In multivariate regression models, self-efficacy was positively associated with scores on the total TIP-RFT and on the Education/Vocation Planning and Independent Living Skills scales. Older age was independently associated with higher scores on the Independent Living Skills scale and higher stress levels were independently associated with lower scores on Education/Vocation Planning scale. CONCLUSION: The TIP-RFT assessment, along with measures of self-efficacy and stress, appear to be useful measures of overall transition readiness for youth and young adults with SCD. Future studies should evaluate whether self-management skill development and health outcomes are indeed affected by programs to improve readiness for transition from pediatric to adult care.