PAX6 suppression of glioma angiogenesis and the expression of vascular endothelial growth factor A

We reported that PAX6 suppresses glioblastoma cell growth in vivo and anchorage-independent growth without significant alteration of cell proliferation in vitro, suggesting that PAX6 may alter the tumor microenvironment. Because we found that PAX6 downregulates expression of the gene encoding vascular endothelial growth factor A (VEGFA) in glioma cells, we used a subcutaneous xenograft model to verify PAX6 suppression of VEGFA-induced angiogenesis based on CD31-immunostaining of endothelial cells. The results showed a significant reduction of VEGFA at the transcription level in PAX6-transfected cells in xenografts and PAX6 has a suppressive effect on the microvascular amplification typically seen in glioblastoma. We showed that PAX6 suppression of VEGFA expression requires its DNA binding-domain. The C-terminal truncation mutant of PAX6, however, did not show the dominant negative function in regulating VEGFA expression that it showed previously in regulating MMP2 expression. In the glioma cell line U251HF, we further determined that blocking the PI3K/Akt signaling pathway with either adenoviral-mediated PTEN expression or LY294002 enhanced PAX6-mediated suppression of VEGFA in an additive manner; thus, PAX6-mediated suppression of VEGFA is not via the canonical pathway through HIF1A. These two VEGFA-regulatory pathways can also be similarly modulated in another malignant glioma cell line, U87, but not in LN229 where the basal VEGFA level is low and PTEN is wild-type. PAX6 suppression of VEGFA appears to be blocked in LN229. In conclusion, our data showed that PAX6 can initiate in glioma cells a new signaling pathway independent of PI3K/Akt-HIF1A signaling to suppress VEGFA expression

Primary leptomeningeal oligodendrogliomatosis

Primary leptomeningeal oligodendrogliomas (PLOs) are rare intracranial malignancies where tumors grow in the subarachnoid space without an obvious connection to the brain or spinal cord parenchyma. Adding to the three previously reported cases of PLO with no parenchymal involvement we report a fourth case of the same in this paper in a 50-year-old woman presenting with unrelenting headaches. CT scan of her head revealed hydrocephalus and MRI revealed diffuse enhancement of her leptomeninges throughout her brain and spine, prominent over the basilar region. Biopsy obtained using a frameless stereotactic biopsy showed sharply defined cell borders, clear cytoplasm, and rounded nuclei consistent with an oligodendroglioma. Our case suggests that PLO can mimic diffuse forms of granulomatous meningitis and should be suspected in patients that clinically and radiographically present like granulomatous meningitis but without blood or CSF markers for the same

EFEMP1 suppresses malignant glioma growth and exerts its action within the tumor extracellular compartment

<p>Abstract</p> <p>Purpose</p> <p>There are conflicting reports regarding the function of EFEMP1 in different cancer types. In this study, we sought to evaluate the role of EFEMP1 in malignant glioma biology.</p> <p>Experimental Design</p> <p>Real-time qRT-PCR was used to quantify <it>EFEMP1 </it>expression in 95 glioblastoma multiforme (GBM). Human high-grade glioma cell lines and primary cultures were engineered to express ectopic EFEMP1, a small hairpin RNA of EFEMP1, or treated with exogenous recombinant EFEMP1 protein. Following treatment, growth was assayed both <it>in vitro </it>and <it>in vivo </it>(subcutaneous (s.c.) and intracranial (i.c.) xenograft model systems).</p> <p>Results</p> <p>Cox regression revealed that EFEMP1 is a favorable prognostic marker for patients with GBM. Over-expression of EFEMP1 eliminated tumor development and suppressed angiogenesis, cell proliferation, and VEGFA expression, while the converse was true with knock-down of endogenous EFEMP1 expression. The EFEMP1 suppression of tumor onset time was nearly restored by ectopic VEGFA expression; however, overall tumor growth rate remained suppressed. This suggested that inhibition of angiogenesis was only partly responsible for EFEMP1's impact on glioma development. In glioma cells that were treated by exogenous EFEMP1 protein or over-expressed endogenous EFEMP1, the EGFR level was reduced and AKT signaling activity attenuated. Mixing of EFEMP1 protein with cells prior to s.c. and i.c. implantations or injection of the protein around the established s.c. xenografts, both significantly suppressed tumorigenicity.</p> <p>Conclusions</p> <p>Overall, our data reveals that EEFEMP1 suppresses glioma growth <it>in vivo</it>, both by modulating the tumor extracellular microenvironment and by altering critical intracellular oncogenic signaling pathways.</p

Pitfalls in assessing stromal tumor infiltrating lymphocytes (sTILs) in breast cancer

Abstract: Stromal tumor-infiltrating lymphocytes (sTILs) are important prognostic and predictive biomarkers in triple-negative (TNBC) and HER2-positive breast cancer. Incorporating sTILs into clinical practice necessitates reproducible assessment. Previously developed standardized scoring guidelines have been widely embraced by the clinical and research communities. We evaluated sources of variability in sTIL assessment by pathologists in three previous sTIL ring studies. We identify common challenges and evaluate impact of discrepancies on outcome estimates in early TNBC using a newly-developed prognostic tool. Discordant sTIL assessment is driven by heterogeneity in lymphocyte distribution. Additional factors include: technical slide-related issues; scoring outside the tumor boundary; tumors with minimal assessable stroma; including lymphocytes associated with other structures; and including other inflammatory cells. Small variations in sTIL assessment modestly alter risk estimation in early TNBC but have the potential to affect treatment selection if cutpoints are employed. Scoring and averaging multiple areas, as well as use of reference images, improve consistency of sTIL evaluation. Moreover, to assist in avoiding the pitfalls identified in this analysis, we developed an educational resource available at www.tilsinbreastcancer.org/pitfalls

Report on computational assessment of Tumor Infiltrating Lymphocytes from the International Immuno-Oncology Biomarker Working Group

Funder: U.S. Department of Health & Human Services | NIH | National Cancer Institute (NCI)Funder: National Center for Research Resources under award number 1 C06 RR12463-01, VA Merit Review Award IBX004121A from the United States Department of Veterans Affairs Biomedical Laboratory Research and Development Service, the DOD Prostate Cancer Idea Development Award (W81XWH-15-1-0558), the DOD Lung Cancer Investigator-Initiated Translational Research Award (W81XWH-18-1-0440), the DOD Peer Reviewed Cancer Research Program (W81XWH-16-1-0329), the Ohio Third Frontier Technology Validation Fund, the Wallace H. Coulter Foundation Program in the Department of Biomedical Engineering and the Clinical and Translational Science Award Program (CTSA) at Case Western Reserve University.Funder: Susan G Komen Foundation (CCR CCR18547966) and a Young Investigator Grant from the Breast Cancer Alliance.Funder: The Canadian Cancer SocietyFunder: Breast Cancer Research Foundation (BCRF), Grant No. 17-194Abstract: Assessment of tumor-infiltrating lymphocytes (TILs) is increasingly recognized as an integral part of the prognostic workflow in triple-negative (TNBC) and HER2-positive breast cancer, as well as many other solid tumors. This recognition has come about thanks to standardized visual reporting guidelines, which helped to reduce inter-reader variability. Now, there are ripe opportunities to employ computational methods that extract spatio-morphologic predictive features, enabling computer-aided diagnostics. We detail the benefits of computational TILs assessment, the readiness of TILs scoring for computational assessment, and outline considerations for overcoming key barriers to clinical translation in this arena. Specifically, we discuss: 1. ensuring computational workflows closely capture visual guidelines and standards; 2. challenges and thoughts standards for assessment of algorithms including training, preanalytical, analytical, and clinical validation; 3. perspectives on how to realize the potential of machine learning models and to overcome the perceptual and practical limits of visual scoring

Application of a risk-management framework for integration of stromal tumor-infiltrating lymphocytes in clinical trials

Funder: Breast Cancer Research Foundation (BCRF); doi: https://doi.org/10.13039/100001006Abstract: Stromal tumor-infiltrating lymphocytes (sTILs) are a potential predictive biomarker for immunotherapy response in metastatic triple-negative breast cancer (TNBC). To incorporate sTILs into clinical trials and diagnostics, reliable assessment is essential. In this review, we propose a new concept, namely the implementation of a risk-management framework that enables the use of sTILs as a stratification factor in clinical trials. We present the design of a biomarker risk-mitigation workflow that can be applied to any biomarker incorporation in clinical trials. We demonstrate the implementation of this concept using sTILs as an integral biomarker in a single-center phase II immunotherapy trial for metastatic TNBC (TONIC trial, NCT02499367), using this workflow to mitigate risks of suboptimal inclusion of sTILs in this specific trial. In this review, we demonstrate that a web-based scoring platform can mitigate potential risk factors when including sTILs in clinical trials, and we argue that this framework can be applied for any future biomarker-driven clinical trial setting

Primary leptomeningeal oligodendrogliomatosis

Primary leptomeningeal oligodendrogliomas (PLOs) are rare intracranial malignancies where tumors grow in the subarachnoid space without an obvious connection to the brain or spinal cord parenchyma. Adding to the three previously reported cases of PLO with no parenchymal involvement we report a fourth case of the same in this paper in a 50-year-old woman presenting with unrelenting headaches. CT scan of her head revealed hydrocephalus and MRI revealed diffuse enhancement of her leptomeninges throughout her brain and spine, prominent over the basilar region. Biopsy obtained using a frameless stereotactic biopsy showed sharply defined cell borders, clear cytoplasm, and rounded nuclei consistent with an oligodendroglioma. Our case suggests that PLO can mimic diffuse forms of granulomatous meningitis and should be suspected in patients that clinically and radiographically present like granulomatous meningitis but without blood or CSF markers for the same

Primary leptomeningeal plasmablastic lymphoma.

Lymphomas that develop in human immunodeficiency virus (HIV) infected patients are predominantly aggressive B-cells lymphomas. The most common HIV-associated lymphomas include Burkitt lymphoma, diffuse large B-cell lymphoma (that often involves the CNS), primary effusion lymphoma, and plasmablastic lymphoma (PBL). Of these, PBL is relatively uncommon and displays a distinct affinity for presentation in the oral cavity. In this manuscript we report a previously undescribed primary leptomeningeal form of PBL in a patient with acquired immunodeficiency syndrome. A 40-year-old HIV positive man presented with acute onset confusion, emesis, and altered mental status. Lumbar puncture showed numerous nucleated cells with atypical plasmocyte predominance. CSF flowcytometry showed kappa restriction with CD8 and CD38 positivity and negative lymphocyte markers, while the MRI showed diffuse leptomeningeal enhancement. As the extensive systemic work-up failed to reveal any disease outside the brain, an en bloc diagnostic brain and meningeal biopsy was performed. The biopsy specimen showed sheets of plasmacytoid cells with one or more large nuclei, prominent nuclear chromatin, scattered mitoses, and abundant cytoplasm, highly suggestive of plasmablastic lymphoma. HIV-associated malignancies have protean and often confusing presentations, which pose diagnostic difficulties posed to the practicing neurological-surgeons. Even in cases where an infectious cause is suspected for the meningeal enhancement, neoplastic involvement should be considered, and cytology and flow-cytometry should be routinely ordered on the CSF samples