Safety and Feasibility of Stereotactic Radiosurgery for Patients with 15 or more Brain Metastases.

Background: Current standard of care treatment for patients with ≥15 brain metastases (BM) is whole brain radiation therapy (WBRT), despite poor neurocognitive outcomes. We analyzed our institutional experience of treating these patients with stereotactic radiosurgery (SRS), with the aim of evaluating safety, cognitive outcomes, and survival metrics. Methods: Patients who received SRS for ≥15 BMs in 1 to 5 fractions from 2014 to 2022 were included. Cognitive outcomes were objectively evaluated using serial Patient-Reported Outcome Measurement Information System (PROMIS) scores. The Kaplan-Meier method was used for survival analysis and log-rank test for intergroup comparisons. Results: Overall, 118 patients underwent 124 courses of LINAC-based SRS. The median number of lesions treated per course was 20 (range, 15-94). Most patients received fractionated SRS to a dose of 24 Gy in 3 fractions (81.5%). At the time of SRS, 19.4% patients had received prior WBRT, and 24.2% had received prior SRS. The rate of any grade radiation necrosis (RN) and grade ≥3 RN were 15.3% and 3.2%, respectively. When evaluating longitudinal PROMIS score trends, 25 of 31 patients had a stable/improved PROMIS score. Patients who did not receive prior brain RT had a longer median survival (7.4 months vs 4.6 months, P = .034). The 12m local control was 97.6%, and the cumulative incidence of distant intracranial failure, with death as a competing event, was 46% (95% CI, 36%, 55%). One year freedom from neurologic death, leptomeningeal disease, and salvage WBRT were 89%, 94.6%, and 84%, respectively. Conclusion: We present here one of the largest studies evaluating SRS for patients with ≥15 BMs. SRS was safe, had favorable cognitive outcomes, and had comparable survival outcomes to contemporary studies evaluating WBRT in this population. Treatment-naïve patients had a median survival of >6 months, long enough to benefit from cognitive sparing with SRS. Our study supports randomized studies comparing SRS and hippocampal avoidance WBRT approaches for these patients.

Fractionated pre-operative stereotactic radiotherapy for patients with brain metastases: a multi-institutional analysis.

BACKGROUND: The current standard of care for patients with a large brain metastasis and limited intracranial disease burden is surgical resection and post-operative single fraction stereotactic radiosurgery (SRS). However, post-operative SRS can still lead to substantial rates of local failure (LF), radiation necrosis (RN), and meningeal disease (MD). Pre-operative SRS may reduce the risk of RN and MD, while fractionated treatments may improve local control by allowing delivery of higher biological effective dose. We hypothesize that pre-operative fractionated stereotactic radiation therapy (FSRT) can minimize rates of LF, RN, and MD. METHODS: A retrospective, multi-institutional analysis was conducted and included patients who had pre-operative FSRT for a large or symptomatic brain metastasis. Pertinent demographic, clinical, radiation, surgical, and follow up data were collected for each patient. A primary measurement was the rate of a composite endpoint of (1) LF, (2) MD, and/or (3) Grade 2 or higher (symptomatic) RN. RESULTS: 53 patients with 55 lesions were eligible for analysis. FSRT was prescribed to a dose of 24-25 Gy in 3-5 fractions. There were 0 LFs, 3 Grade 2-3 RN events, and 1 MD occurrence, which corresponded to an 8% per-patient composite endpoint event rate. CONCLUSIONS: In this study, the composite endpoint of 8% for pre-operative FSRT was improved compared to previously reported rates with post-operative SRS of 49-60% (N107C, Mahajan etal. JCOG0504) and pre-operative SRS endpoints of 20.6% (PROPS-BM). Pre-operative FSRT appears to be safe, effective, and may decrease the incidence of adverse outcomes. Prospective validation is needed.

Cerebellopontine angle ependymoma presenting as isolated hearing loss in an elderly patient: A case report and literature review.

BACKGROUND: Ependymoma is an uncommon tumor accounting for approximately 1.9% of all adult central nervous system tumors. Ependymomas at the cerebellopontine angle (CPA) are even more rare and only previously described in isolated case reports. Typically, acoustic neuromas and meningiomas represent the bulk of adult CPA tumors. Diagnosis can be challenging, as ependymomas have clinical findings and imaging characteristics that overlap with more common tumor histologies at the CPA. CASE DESCRIPTION: We present the case of a 70-year-old male patient with progressive, isolated left-sided hearing loss found to have a World Health Organization (WHO) Grade II CPA ependymoma, representing one of the oldest recorded patients presenting with this primarily pediatric malignancy in this unique location. The patient presentation with isolated hearing loss was particularly unusual. When associated with neurologic deficits, CPA ependymomas more characteristically result in facial nerve impairment with fully preserved hearing, while vestibular schwannomas tend to present with isolated hearing loss. The standard of care for pediatric ependymomas is maximal safe resection with adjuvant radiotherapy, but treatment paradigms in adult CPA ependymoma are not well defined particularly for WHO Grade II disease. After resection, he received adjuvant radiation to decrease the risk of local recurrence. Twenty-nine months after resection, the patient remains free of treatment-related toxicity or disease recurrence. CONCLUSION: We review this patient's clinical course in the context of the literature to highlight the challenges associated with timely diagnosis of this rare tumor and the controversial role of adjuvant therapy in preventing local recurrence in these patients.

Oncolytic herpes simplex virus infects myeloma cells in vitro and in vivo.

Because most patients with multiple myeloma (MM) develop resistance to current regimens, novel approaches are needed. Genetically modified, replication-competent oncolytic viruses exhibit high tropism for tumor cells regardless of cancer stage and prior treatment. Receptors of oncolytic herpes simplex virus 1 (oHSV-1), NECTIN-1, and HVEM are expressed on MM cells, prompting us to investigate the use of oHSV-1 against MM. Using oHSV-1-expressing GFP, we found a dose-dependent increase in the GFP+ signal in MM cell lines and primary MM cells. Whereas NECTIN-1 expression is variable among MM cells, we discovered that HVEM is ubiquitously and highly expressed on all samples tested. Expression of HVEM was consistently higher on CD138+/CD38+ plasma cells than in non-plasma cells. HVEM blocking demonstrated the requirement of this receptor for infection. However, we observed that, although oHSV-1 could efficiently infect and kill all MM cell lines tested, no viral replication occurred. Instead, we identified that oHSV-1 induced MM cell apoptosis via caspase-3 cleavage. We further noted that oHSV-1 yielded a significant decrease in tumor volume in two mouse xenograft models. Therefore, oHSV-1 warrants exploration as a novel potentially effective treatment option in MM, and HVEM should be investigated as a possible therapeutic target.

Malignant ossifying fibromyxoid tumor of the brain treated with post-operative fractionated stereotactic radiation therapy: A case report and literature review.

BACKGROUND: Ossifying fibromyxoid tumor (OFMT) is a rare musculoskeletal soft-tissue neoplasm of uncertain histogenesis most frequently occurring in the lower extremities. Conventionally, considered benign, these tumors are often managed by surgical resection followed by surveillance. However, malignant OFMTs with an increased propensity for local recurrence and distant metastasis have been recently identified, and the role of adjuvant therapy in these more aggressive cases is unclear. CASE DESCRIPTION: We present, to the best of our knowledge, the first reported case of a primary, malignant, and intracranial OFMT. A 29-year-old female presented with recurrent headaches secondary to a large mass in her right frontal lobe. She underwent gross total resection of the brain mass with final pathology consistent with malignant OFMT demonstrating high-risk features including increased cellularity, grade, and mitotic activity. Due to these high-risk features, she received postoperative fractionated stereotactic radiation therapy (FSRT) to the resection cavity, and to the best of our knowledge, she represents the only known patient with OFMT to be treated with adjuvant FSRT. She tolerated the adjuvant treatment well with no acute or late toxicities and remains disease-free over 5 ½ years after resection. CONCLUSION: Adjuvant FSRT appears to be a safe and efficacious approach for managing this rare intracranial disease presentation. We review this patient's clinical course in the context of the literature to demonstrate the difficulties associated with accurate diagnosis of this rare tumor and the controversial role of adjuvant therapy in preventing disease recurrence in this patient population.

MiR-16 regulates crosstalk in NF-κB tolerogenic inflammatory signaling between myeloma cells and bone marrow macrophages.

High levels of circulating miR-16 in the serum of multiple myeloma (MM) patients are independently associated with longer survival. Although the tumor suppressor function of intracellular miR-16 in MM plasma cells (PCs) has been elucidated, its extracellular role in maintaining a nonsupportive cancer microenvironment has not been fully explored. Here, we show that miR-16 is abundantly released by MM cells through extracellular vesicles (EVs) and that differences in its intracellular expression as associated with chromosome 13 deletion (Del13) are correlated to extracellular miR-16 levels. We also demonstrate that EVs isolated from MM patients and from the conditioned media of MM-PCs carrying Del13 more strongly differentiate circulating monocytes to M2-tumor supportive macrophages (TAMs), compared with MM-PCs without this chromosomal aberration. Mechanistically, our data show that miR-16 directly targets the IKKα/ß complex of the NF-κB canonical pathway, which is critical not only in supporting MM cell growth, but also in polarizing macrophages toward an M2 phenotype. By using a miR-15a-16-1-KO mouse model, we found that loss of the miR-16 cluster supports polarization to M2 macrophages. Finally, we demonstrate the therapeutic benefit of miR-16 overexpression in potentiating the anti-MM activity by a proteasome inhibitor in the presence of MM-resident bone marrow TAM.

Daratumumab induces CD38 internalization and impairs myeloma cell adhesion.

Daratumumab (Dara), a human immunoglobulin G1 kappa (IgG1κ) monoclonal anti-CD38 antibody, has been approved by the U.S. Food and Drug Administration for the treatment of relapsed multiple myeloma (MM) as a single agent as well as in combination with immunomodulatory drugs (IMiDs) and proteasome inhibitors (PI). Although the scientific rationale behind the use of Dara in combination with IMiDs has been extensively explored, the molecular mechanisms underlying Dara-PI regimens have not yet been investigated. Here, we demonstrate that CD38 on the surface of MM cells is rapidly internalized after Dara treatment; we also show that Dara treatment impairs MM cell adhesion, an effect that can be rescued by using the endocytosis inhibitor Dynasore. Finally, we show that Dara potentiates bortezomib (BTZ) killing of MM cells in vitro and in vivo, independent of its function as an immune activator. In conclusion, our data show that Dara impairs MM cell adhesion, which results in an increased sensitivity of MM to proteasome inhibition.

Transcriptional regulation of microRNA-100, -146a, and -150 genes by p53 and NFκB p65/RelA in mouse striatal STHdh(Q7)/ Hdh(Q7) cells and human cervical carcinoma HeLa cells.

MicroRNA (miRNA) genes generally share many features common to those of protein coding genes. Various transcription factors (TFs) and co-regulators are also known to regulate miRNA genes. Here we identify novel p53 and NFκB p65/RelA responsive miRNAs and demonstrate that these 2 TFs bind to the regulatory sequences of miR-100, -146a and -150 in both mouse striatal and human cervical carcinoma cells and regulate their expression. p53 represses the miRNAs while NFκB p65/RelA induces them. Further, we provide evidence that exogenous p53 inhibits NFκB p65/RelA activity by reducing its nuclear content and competing with it for CBP binding. This suggests for the existence of a functional cross-talk between the 2 TFs in regulating miRNA expression. Moreover, promoter occupancy assay reveals that exogenous p53 excludes NFκB p65/RelA from its binding site in the upstream sequence of miR-100 gene thereby causing its repression. Thus, our work identifies novel p53 and NFκB p65/RelA responsive miRNAs in human and mouse and uncovers possible mechanisms of co-regulation of miR-100. It is to be mentioned here that cross-talks between p53 and NFκB p65/RelA have been observed to define the outcome of several biological processes and that the pro-apoptotic effect of p53 and the pro-survival functions of NFκB can be largely mediated via the biological roles of the miRNAs these TFs regulate. Our observation with cell lines thus provides an important platform upon which further work is to be done to establish the biological significance of such co-regulation of miRNAs by p53 and NFκB p65/RelA.

Micro RNA -214,-150,-146a and-125b target Huntingtin gene.

We observed earlier that expressions of several micro RNAs (miRNAs) are altered in a cell model of Huntington's disease (HD). As genes involved in different neurodegenerative diseases are targeted by miRNAs, we searched databases to find out whether Huntingtin gene (HTT), mutation to which causes HD, is a target of any miRNA. Among many miRNAs that are predicted to target HTT, we showed using various experimental approaches that miR-214, miR-150, miR-146a and miR-125b could target both human HTT and mouse Htt. Luciferase reporter vectors containing 3'-UTRs of mouse and human HTT showed reduction in relative luciferase activities on exogenous expression of cloned miRNAs.  Loss of function studies with miRNA inhibitors led to the revival of luciferase activities of cloned 3'-UTR of HTT.  Exogenous expression of these miRNAs reduced the expression of endogenous Htt in mouse cells. Expression of miRNAs with mutations at seed sequences neither reduced the reporter luciferase activities nor the endogenous expression of Htt. Taking together, we showed that miR-214, miR-150, miR-146a and miR-125b targeted HTT. Besides, the exogenous expression of wild type miRNAs reduced HTT aggregates formed by the recombinant exon1 of HTT gene that codes for 83Q tagged with 3'-UTR of Htt, as observed by filter retardation assay and confocal microscopy. In summary, regulation of HTT by miRNAs might provide a new mechanism for the modulation of HD. 

Regulation of miR-146a by RelA/NFkB and p53 in STHdh(Q111)/Hdh(Q111) cells, a cell model of Huntington's disease.

Huntington's disease (HD) is caused by the expansion of N-terminal polymorphic poly Q stretch of the protein huntingtin (HTT). Deregulated microRNAs and loss of function of transcription factors recruited to mutant HTT aggregates could cause characteristic transcriptional deregulation associated with HD. We observed earlier that expressions of miR-125b, miR-146a and miR-150 are decreased in STHdh(Q111)/Hdh(Q111) cells, a model for HD in comparison to those of wild type STHdh(Q7)/Hdh(Q7) cells. In the present manuscript, we show by luciferase reporter assays and real time PCR that decreased miR-146a expression in STHdh(Q111)/Hdh(Q111) cells is due to decreased expression and activity of p65 subunit of NFkB (RelA/NFkB). By reporter luciferase assay, RT-PCR and western blot analysis, we also show that both miR-150 and miR-125b target p53. This partially explains the up regulation of p53 observed in HD. Elevated p53 interacts with RelA/NFkB, reduces its expression and activity and decreases the expression of miR-146a, while knocking down p53 increases RelA/NFkB and miR-146a expressions. We also demonstrate that expression of p53 is increased and levels of RelA/NFkB, miR-146a, miR-150 and miR-125b are decreased in striatum of R6/2 mice, a mouse model of HD and in cell models of HD. In a cell model, this effect could be reversed by exogenous expression of chaperone like proteins HYPK and Hsp70. We conclude that (i) miR-125b and miR-150 target p53, which in turn regulates RelA/NFkB and miR-146a expressions; (ii) reduced miR-125b and miR-150 expressions, increased p53 level and decreased RelA/NFkB and miR-146a expressions originate from mutant HTT (iii) p53 directly or indirectly regulates the expression of miR-146a. Our observation of interplay between transcription factors and miRNAs using HD cell model provides an important platform upon which further work is to be done to establish if such regulation plays any role in HD pathogenesis.

Altered microRNAs in STHdh(Q111)/Hdh(Q111) cells: miR-146a targets TBP.

We studied expression of 90 miRNAs in STHdh(Q111)/Hdh(Q111) cells, a model for Huntington's disease and compared with that obtained in STHdh(Q7)/Hdh(Q7) cells. Fifteen miRNAs were down regulated and 12 miRNAs were up regulated more than 2-fold. Such changes were statistically significant. One hundred and forty-two genes are experimentally known targets of these altered miRNAs. It has been predicted that miR-146a may target Tata Binding Protein (TBP). Using luciferase reporter assays with 3'-UTRs of TBP, over-expression and inhibition of miR-146a, we showed that miR-146a targets TBP. Regulation of TBP by miR-146a may contribute to HD pathogenesis.

Immune responses in kala-azar.

Human infection with Leishmania results in diverse clinical and immunopathological situations. The capacity of the parasites to cause this wide range of disease manifestations depends upon their ability to evade the immune defense mechanisms by performing a well-tuned orchestra of hostparasite interactions inside the macrophages. While updated knowledge focus on the key role of cell-mediated immunity (CMI) in protection, the survival strategies of the parasites leads to the suppression of CMI which can further be aggravated by the co-infections with HIV, tuberculosis etc. The present review describes the immune mechanisms in human leishmaniasis with a special attention to visceral leishmaniasis or kala-azar, one of the most important epidemiological health problems in Indian subcontinent. Modulations of the both humoral and cell-mediated immune responses during asymptomatic infections, active disease and after successful chemotherapy are discussed. The components responsible for the regulation of the critical balance of Th1/Th2 type of responses are re-evaluated. Co-infection of HIV and visceral leishmaniasis and their interdependence has been addressed. Although the specific role of an elevated humoral response in kala-azar is yet to be established, attempts for its application in diagnosis, precisely for the development of field diagnostic techniques, are presented. Also discussed are attempts to utilize the immunogenic potentials of different leishmanial antigens in the development of anti-leishmanial vaccines.