Magnetic Resonance Imaging of Iron Metabolism with T2* Mapping Predicts an Enhanced Clinical Response to Pharmacologic Ascorbate in Patients with GBM.

PURPOSE: Pharmacologic ascorbate (P-AscH-) is hypothesized to be an iron (Fe)-dependent tumor-specific adjuvant to chemoradiation in treating glioblastoma (GBM). This study determined the efficacy of combining P-AscH- with radiation and temozolomide in a phase II clinical trial while simultaneously investigating a mechanism-based, noninvasive biomarker in T2* mapping to predict GBM response to P-AscH- in humans. PATIENTS AND METHODS: The single-arm phase II clinical trial (NCT02344355) enrolled 55 subjects, with analysis performed 12 months following the completion of treatment. Overall survival (OS) and progression-free survival (PFS) were estimated with the Kaplan-Meier method and compared across patient subgroups with log-rank tests. Forty-nine of 55 subjects were evaluated using T2*-based MRI to assess its utility as an Fe-dependent biomarker. RESULTS: Median OS was estimated to be 19.6 months [90% confidence interval (CI), 15.7-26.5 months], a statistically significant increase compared with historic control patients (14.6 months). Subjects with initial T2* relaxation < 50 ms were associated with a significant increase in PFS compared with T2*-high subjects (11.2 months vs. 5.7 months, P < 0.05) and a trend toward increased OS (26.5 months vs. 17.5 months). These results were validated in preclinical in vitro and in vivo model systems. CONCLUSIONS: P-AscH- combined with temozolomide and radiotherapy has the potential to significantly enhance GBM survival. T2*-based MRI assessment of tumor iron content is a prognostic biomarker for GBM clinical outcomes. See related commentary by Nabavizadeh and Bagley, p. 255.

Targeting the Retinoid X Receptor Pathway Prevents and Ameliorates Murine Chronic Graft-Versus-Host Disease.

Most allogeneic hematopoietic stem cell transplant (allo-HSCT) recipients receive peripheral blood stem cell grafts resulting in a 30%-70% incidence of chronic graft-versus-host disease (cGVHD), a major cause of mortality and morbidity in long-term survivors. While systemic steroids remain the standard of care for first-line therapy, patients may require long-term administration, and those with steroid-resistant or refractory cGVHD have a worse prognosis. Although durable and deep responses with second-line therapies can be achieved in some patients, there remains an urgent need for new therapies. In this study, we evaluated the efficacy of IRX4204, a novel agonist that activates RXRs and is in clinical trials for cancer treatment to prevent and treat cGVHD in two complementary murine models. In a major histocompatibility complex mismatched, non-sclerodermatous multiorgan system model with bronchiolitis obliterans, IRX4204 prevented and reversed cGVHD including associated pulmonary dysfunction with restoration of germinal center T-follicular helper: T-follicular regulatory cell balance. In a minor histocompatibility antigen disparate sclerodermatous model, IRX4204 treatment significantly prevented and ameliorated skin cGVHD by reducing Th1 and Th17 differentiation due to anti-inflammatory properties. Together, these results indicate that IRX4204 is a promising therapeutic option to treat cGVHD with bronchiolitis obliterans or sclerodermatous manifestations.

RNA helicase DDX21 mediates nucleotide stress responses in neural crest and melanoma cells.

The availability of nucleotides has a direct impact on transcription. The inhibition of dihydroorotate dehydrogenase (DHODH) with leflunomide impacts nucleotide pools by reducing pyrimidine levels. Leflunomide abrogates the effective transcription elongation of genes required for neural crest development and melanoma growth in vivo1. To define the mechanism of action, we undertook an in vivo chemical suppressor screen for restoration of neural crest after leflunomide treatment. Surprisingly, we found that alterations in progesterone and progesterone receptor (Pgr) signalling strongly suppressed leflunomide-mediated neural crest effects in zebrafish. In addition, progesterone bypasses the transcriptional elongation block resulting from Paf complex deficiency, rescuing neural crest defects in ctr9 morphant and paf1(alnz24) mutant embryos. Using proteomics, we found that Pgr binds the RNA helicase protein Ddx21. ddx21-deficient zebrafish show resistance to leflunomide-induced stress. At a molecular level, nucleotide depletion reduced the chromatin occupancy of DDX21 in human A375 melanoma cells. Nucleotide supplementation reversed the gene expression signature and DDX21 occupancy changes prompted by leflunomide. Together, our results show that DDX21 acts as a sensor and mediator of transcription during nucleotide stress.

An integrated physico-chemical approach for explaining the differential impact of FLASH versus conventional dose rate irradiation on cancer and normal tissue responses.

For decades the field of radiation oncology has sought to improve the therapeutic ratio through innovations in physics, chemistry, and biology. To date, technological advancements in image guided beam delivery techniques have provided clinicians with their best options for improving this critical tool in cancer care. Medical physics has focused on the preferential targeting of tumors while minimizing the collateral dose to the surrounding normal tissues, yielding only incremental progress. However, recent developments involving ultra-high dose rate irradiation termed FLASH radiotherapy (FLASH-RT), that were initiated nearly 50 years ago, have stimulated a renaissance in the field of radiotherapy, long awaiting a breakthrough modality able to enhance therapeutic responses and limit normal tissue injury. Compared to conventional dose rates used clinically (0.1-0.2 Gy/s), FLASH can implement dose rates of electrons or X-rays in excess of 100 Gy/s. The implications of this ultra-fast delivery of dose are significant and need to be re-evaluated to appreciate the fundamental aspects underlying this seemingly unique radiobiology. The capability of FLASH to significantly spare normal tissue complications in multiple animal models, when compared to conventional rates of dose-delivery, while maintaining persistent growth inhibition of select tumor models has generated considerable excitement, as well as skepticism. Based on fundamental principles of radiation physics, radio-chemistry, and tumor vs. normal cell redox metabolism, this article presents a series of testable, biologically relevant hypotheses, which may help rationalize the differential effects of FLASH irradiation observed between normal tissue and tumors.

Notch signaling mediated by Delta-like ligands 1 and 4 controls the pathogenesis of chronic GVHD in mice.

Chronic graft-versus-host disease (cGVHD) is a major complication of allogeneic hematopoietic cell transplantation (allo-HCT) and remains an area of unmet clinical need with few treatment options available. Notch blockade prevents acute GVHD in multiple mouse models, but the impact of Notch signaling on cGVHD remains unknown. Using genetic and antibody-mediated strategies of Notch inhibition, we investigated the role of Notch signaling in complementary mouse cGVHD models that mimic several aspects of human cGVHD in search of candidate therapeutics. In the B10.D2→BALB/c model of sclerodermatous cGVHD, Delta-like ligand 4 (Dll4)-driven Notch signaling was essential for disease development. Antibody-mediated Dll4 inhibition conferred maximum benefits when pursued early in a preventative fashion, with anti-Dll1 enhancing early protection. Notch-deficient alloantigen-specific T cells showed no early defects in proliferation or helper polarization in vivo but subsequently exhibited markedly decreased cytokine secretion and enhanced accumulation of FoxP3+ regulatory T cells. In the B6→B10.BR major histocompatibility complex-mismatched model with multi-organ system cGVHD and prominent bronchiolitis obliterans (BO), but not skin manifestations, absence of Notch signaling in T cells provided long-lasting disease protection that was replicated by systemic targeting of Dll1, Dll4, or both Notch ligands, even during established disease. Notch inhibition decreased target organ damage and germinal center formation. Moreover, decreased BO-cGVHD was observed upon inactivation of Notch1 and/or Notch2 in T cells. Systemic targeting of Notch2 alone was safe and conferred therapeutic benefits. Altogether, Notch ligands and receptors regulate key pathogenic steps in cGVHD and emerge as novel druggable targets to prevent or treat different forms of cGVHD.

Targeted Rho-associated kinase 2 inhibition suppresses murine and human chronic GVHD through a Stat3-dependent mechanism.

Chronic graft-versus-host disease (cGVHD) remains a major complication following allogeneic bone marrow transplantation (BMT). The discovery of novel therapeutics is dependent on assessment in preclinical murine models of cGVHD. Rho-associated kinase 2 (ROCK2) recently was shown to be implicated in regulation of interleukin-21 (IL-21) and IL-17 secretion in mice and humans. Here, we report that the selective ROCK2 inhibitor KD025 effectively ameliorates cGVHD in multiple models: a full major histocompatibility complex (MHC) mismatch model of multiorgan system cGVHD with bronchiolitis obliterans syndrome and a minor MHC mismatch model of sclerodermatous GVHD. Treatment with KD025 resulted in normalization of pathogenic pulmonary function, which correlates with a marked reduction of antibody and collagen deposition in the lungs of treated mice to levels comparable to non-cGVHD controls. Spleens of mice treated with KD025 had decreased frequency of T follicular helper cells and increased frequency of T follicular regulatory cells, accompanied by a reduction in signal transducer and activator of transcription 3 (STAT3) and concurrent increase in STAT5 phosphorylation. The critical role of STAT3 in this cGVHD model was confirmed by data showing that mice transplanted with inducible STAT3-deficient T cells had pulmonary function comparable to the healthy negative controls. The therapeutic potential of targeted ROCK2 inhibition in the clinic was solidified further by human data demonstrating the KD025 inhibits the secretion of IL-21, IL-17, and interferon γ along with decreasing phosphorylated STAT3 and reduced protein expression of interferon regulatory factor 4 and B-cell lymphoma 6 (BCL6) in human peripheral blood mononuclear cells purified from active cGVHD patients. Together these data highlight the potential of targeted ROCK2 inhibition for clinical cGVHD therapy.

Ibrutinib treatment ameliorates murine chronic graft-versus-host disease.

Chronic graft-versus-host disease (cGVHD) is a life-threatening impediment to allogeneic hematopoietic stem cell transplantation, and current therapies do not completely prevent and/or treat cGVHD. CD4+ T cells and B cells mediate cGVHD; therefore, targeting these populations may inhibit cGVHD pathogenesis. Ibrutinib is an FDA-approved irreversible inhibitor of Bruton's tyrosine kinase (BTK) and IL-2 inducible T cell kinase (ITK) that targets Th2 cells and B cells and produces durable remissions in B cell malignancies with minimal toxicity. Here, we evaluated whether ibrutinib could reverse established cGVHD in 2 complementary murine models, a model interrogating T cell-driven sclerodermatous cGVHD and an alloantibody-driven multiorgan system cGVHD model that induces bronchiolar obliterans (BO). In the T cell-mediated sclerodermatous cGVHD model, ibrutinib treatment delayed progression, improved survival, and ameliorated clinical and pathological manifestations. In the alloantibody-driven cGVHD model, ibrutinib treatment restored pulmonary function and reduced germinal center reactions and tissue immunoglobulin deposition. Animals lacking BTK and ITK did not develop cGVHD, indicating that these molecules are critical to cGVHD development. Furthermore, ibrutinib treatment reduced activation of T and B cells from patients with active cGVHD. Our data demonstrate that B cells and T cells drive cGVHD and suggest that ibrutinib has potential as a therapeutic agent, warranting consideration for cGVHD clinical trials.

Interstitial rotating shield brachytherapy for prostate cancer.

PURPOSE: To present a novel needle, catheter, and radiation source system for interstitial rotating shield brachytherapy (I-RSBT) of the prostate. I-RSBT is a promising technique for reducing urethra, rectum, and bladder dose relative to conventional interstitial high-dose-rate brachytherapy (HDR-BT). METHODS: A wire-mounted 62 GBq(153)Gd source is proposed with an encapsulated diameter of 0.59 mm, active diameter of 0.44 mm, and active length of 10 mm. A concept model I-RSBT needle/catheter pair was constructed using concentric 50 and 75 µm thick nickel-titanium alloy (nitinol) tubes. The needle is 16-gauge (1.651 mm) in outer diameter and the catheter contains a 535 µm thick platinum shield. I-RSBT and conventional HDR-BT treatment plans for a prostate cancer patient were generated based on Monte Carlo dose calculations. In order to minimize urethral dose, urethral dose gradient volumes within 0-5 mm of the urethra surface were allowed to receive doses less than the prescribed dose of 100%. RESULTS: The platinum shield reduced the dose rate on the shielded side of the source at 1 cm off-axis to 6.4% of the dose rate on the unshielded side. For the case considered, for the same minimum dose to the hottest 98% of the clinical target volume (D(98%)), I-RSBT reduced urethral D(0.1cc) below that of conventional HDR-BT by 29%, 33%, 38%, and 44% for urethral dose gradient volumes within 0, 1, 3, and 5 mm of the urethra surface, respectively. Percentages are expressed relative to the prescription dose of 100%. For the case considered, for the same urethral dose gradient volumes, rectum D(1cc) was reduced by 7%, 6%, 6%, and 6%, respectively, and bladder D(1cc) was reduced by 4%, 5%, 5%, and 6%, respectively. Treatment time to deliver 20 Gy with I-RSBT was 154 min with ten 62 GBq (153)Gd sources. CONCLUSIONS: For the case considered, the proposed(153)Gd-based I-RSBT system has the potential to lower the urethral dose relative to HDR-BT by 29%-44% if the clinician allows a urethral dose gradient volume of 0-5 mm around the urethra to receive a dose below the prescription. A multisource approach is necessary in order to deliver the proposed (153)Gd-based I-RSBT technique in reasonable treatment times.