HTLV I is a retrovirus that can result in decreased effectiveness of immune response and an increased incidence of coinfections. HTLV I CNS coinfections with rare clinical manifestations that may relate to impaired cellular immunity are presented.
<p>Supplementary Fig. S1. Immune cell-dependent survival differences between males and females in murine glioblastoma models. Supplementary Fig. S2. Mouse syngeneic GBM cell lines do not contain a Y chromosome. Supplementary Fig. S3. Frequencies of tumor-infiltrating immune cells in SB28 model. Supplementary Fig. S4. Sex difference in survival in SB28-OVA model and GL261 model. Supplementary Fig. S5. No sex difference in T cells was observed at an earlier time point. Supplementary Fig. S6. Male T cells are more exhausted in the GL261 model. Supplementary Fig. S7. Phenotype of T cells in the periphery does not replicate the sex differences shown in tumor-infiltrating T cells. Supplementary Fig. S8. Frequencies of exhausted T cell subsets. Supplementary Fig. S9. PD1 blockade enhanced immune responses Supplementary Fig. S10. No significant difference in the immune cell composition of bone marrow chimera mice before tumor implantation. Supplementary Fig. S11. Male and female CD8+ T cells in mixed bone marrow chimera model. Supplementary Fig. S12. Phenotyping tumor-infiltrating CD8+ T cell from GBM patient tumors. Supplementary Fig. S13. scRNA-seq analysis of tumor-infiltrating CD8+ T cells from GBM patients. Supplementary Fig. S14. mRNA expression level of transcription factors in human in vitro exhausted T cells. Supplementary Fig. S15. Expression level of X chromosome inactivation (XCI) genes in exhausted T cells. Supplementary Fig. S16. Inhibition of UTX abrogates sex differences in T cell exhaustion.</p>
<div>Abstract<p>Sex differences in glioblastoma (GBM) incidence and outcome are well recognized, and emerging evidence suggests that these extend to genetic/epigenetic and cellular differences, including immune responses. However, the mechanisms driving immunologic sex differences are not fully understood. Here, we demonstrate that T cells play a critical role in driving GBM sex differences. Male mice exhibited accelerated tumor growth, with decreased frequency and increased exhaustion of CD8<sup>+</sup> T cells in the tumor. Furthermore, a higher frequency of progenitor exhausted T cells was found in males, with improved responsiveness to anti–PD-1 treatment. Moreover, increased T-cell exhaustion was observed in male GBM patients. Bone marrow chimera and adoptive transfer models indicated that T cell–mediated tumor control was predominantly regulated in a cell-intrinsic manner, partially mediated by the X chromosome inactivation escape gene <i>Kdm6a.</i> These findings demonstrate that sex-biased predetermined behavior of T cells is critical for inducing sex differences in GBM progression and immunotherapy response.</p>Significance:<p>Immunotherapies in patients with GBM have been unsuccessful due to a variety of factors, including the highly immunosuppressive tumor microenvironment in GBM. This study demonstrates that sex-biased T-cell behaviors are predominantly intrinsically regulated, further suggesting sex-specific approaches can be leveraged to potentially improve the therapeutic efficacy of immunotherapy in GBM.</p><p><i><a href="https://aacrjournals.org/cancerdiscovery/article/doi/10.1158/2159-8290.CD-23-0727" target="_blank">See related commentary by Alspach, p. 1966.</a></i></p><p><i><a href="https://aacrjournals.org/cancerdiscovery/article/doi/10.1158/2159-8290.CD-13-9-ITI" target="_blank">This article is featured in Selected Articles from This Issue, p. 1949</a></i></p></div>
HIV-1 entry begins with viral envelope glycoprotein gp120 interacting with host-cell CD4 and an entry coreceptor (mainly chemokine receptors CCR5 or CXCR4). Inhibitors of particular coreceptors are being developed in order to exploit this step of cellular infection. However, effectiveness of these drugs requires matching of the administered therapeutic to coreceptor use by the viral variants infecting each patient. Patient viruses may use only CCR5 (R5), only CXCR4 (X4) or both (D/M). Most patients in early disease have R5 variants, with the presence of X4 variants increasing as disease progresses; the infecting subtype also affects the prevalence of X4 variants. Phenotypic, genotypic and clinical trial tests are in use to determine coreceptor utilization by HIV-1 variants, termed tropism, and to predict the response to entry inhibitors. Maraviroc is the only approved entry-coreceptor inhibitor and inhibits CCR5-gp120 interaction. Clinical trials of maraviroc in specific patient subgroups are elucidating the drug's role in contemporary clinical practice. Treatment failure to this and other CCR5 inhibitors has been shown to result from either outgrowth of X4 variants or through resistance mutations leading to R5 variants that are able to enter cells using drug-bound CCR5; thus, new entry inhibitors seek to circumvent this mechanism of resistance.
Abstract Extracellular vesicles (EVs), including exosomes and microvesicles, are 30–800 nm vesicles that are released by most cell types, as biological packages for intercellular communication. Their importance in cancer and inflammation makes EVs and their cargo promising biomarkers of disease and cell-free therapeutic agents. Emerging high-resolution cytometric methods have created a pressing need for efficient fluorescent labeling procedures to visualize and detect EVs. Suitable labels must be bright enough for one EV to be detected without the generation of label-associated artifacts. To identify a strategy that robustly labels individual EVs, we used nanoFACS, a high-resolution flow cytometric method that utilizes light scattering and fluorescence parameters along with sample enumeration, to evaluate various labels. Specifically, we compared lipid-, protein-, and RNA-based staining methods and developed a robust EV staining strategy, with the amine-reactive fluorescent label, 5-(and-6)-Carboxyfluorescein Diacetate Succinimidyl Ester, and size exclusion chromatography to remove unconjugated label. By combining nanoFACS measurements of light scattering and fluorescence, we evaluated the sensitivity and specificity of EV labeling assays in a manner that has not been described for other EV detection methods. Efficient characterization of EVs by nanoFACS paves the way towards further study of EVs and their roles in health and disease.
Acute cauda equina syndrome is an uncommon but significant neurologic presentation due to a variety of underlying diseases. Anatomical compression of nerve roots, usually by a lumbar disk hernia is a common cause in the general population, while inflammatory, neoplastic, and ischemic causes have also been recognized. Among human immunodeficiency virus (HIV) infected patients with acquired immunodeficiency syndrome, infectious causes are encountered more frequently, the most prevalent of which are: cytomegalovirus, herpes simplex virus 1/2, varicella zoster virus, and Mycobacterium tuberculosis infections. Studies of cauda equina syndrome in well-controlled HIV infection are lacking. We describe such a case of cauda equina syndrome in a well-controlled HIV-infected patient, along with a brief review of the literature regarding the syndrome's diagnosis and treatment in individuals with HIV infection. A 36-year-old Greek male, HIV-positive patient presented with perineal and left hemiscrotal numbness, lumbar pain, left-sided sciatica, and urinary incontinence. Magnetic resonance imaging of the patient's lumbar spine revealed intrathecal migration of a fragment from an intervertebral lumbar disk exerting pressure on the cauda equina. A cerebrospinal fluid examination, brain computed tomography scan, spine magnetic resonance imaging, and serological test results were negative for central nervous system infections. Our patient underwent emergency neurosurgical spinal decompression, which resolved most symptoms, except for mild urinary incontinence. Noninfectious etiologies may also cause cauda equina syndrome in HIV-infected individuals, especially in well-controlled disease under antiretroviral therapy. Prompt recognition and treatment of the underlying cause is important to minimize residual symptoms. Targeted antimicrobial chemotherapy is used to treat infectious causes, while prompt surgical decompression is favored for anatomical causes of cauda equina syndrome in the HIV-infected patient.
Despite therapeutic interventions for glioblastoma (GBM), cancer stem cells (CSCs) drive recurrence. The precise mechanisms underlying CSC resistance, namely inhibition of cell death, are unclear. We built on previous observations that the high cell surface expression of junctional adhesion molecule-A drives CSC maintenance and identified downstream signaling networks, including the cysteine protease inhibitor SerpinB3. Using genetic depletion approaches, we found that SerpinB3 is necessary for CSC maintenance, survival, and tumor growth, as well as CSC pathway activation. Knockdown of SerpinB3 also increased apoptosis and susceptibility to radiation therapy. SerpinB3 was essential to buffer cathepsin L-mediated cell death, which was enhanced with radiation. Finally, we found that SerpinB3 knockdown increased the efficacy of radiation in pre-clinical models. Taken together, our findings identify a GBM CSC-specific survival mechanism involving a cysteine protease inhibitor, SerpinB3, and provide a potential target to improve the efficacy of GBM therapies against therapeutically resistant CSCs.
<p>Supplementary Fig. S1. Immune cell-dependent survival differences between males and females in murine glioblastoma models. Supplementary Fig. S2. Mouse syngeneic GBM cell lines do not contain a Y chromosome. Supplementary Fig. S3. Frequencies of tumor-infiltrating immune cells in SB28 model. Supplementary Fig. S4. Sex difference in survival in SB28-OVA model and GL261 model. Supplementary Fig. S5. No sex difference in T cells was observed at an earlier time point. Supplementary Fig. S6. Male T cells are more exhausted in the GL261 model. Supplementary Fig. S7. Phenotype of T cells in the periphery does not replicate the sex differences shown in tumor-infiltrating T cells. Supplementary Fig. S8. Frequencies of exhausted T cell subsets. Supplementary Fig. S9. PD1 blockade enhanced immune responses Supplementary Fig. S10. No significant difference in the immune cell composition of bone marrow chimera mice before tumor implantation. Supplementary Fig. S11. Male and female CD8+ T cells in mixed bone marrow chimera model. Supplementary Fig. S12. Phenotyping tumor-infiltrating CD8+ T cell from GBM patient tumors. Supplementary Fig. S13. scRNA-seq analysis of tumor-infiltrating CD8+ T cells from GBM patients. Supplementary Fig. S14. mRNA expression level of transcription factors in human in vitro exhausted T cells. Supplementary Fig. S15. Expression level of X chromosome inactivation (XCI) genes in exhausted T cells. Supplementary Fig. S16. Inhibition of UTX abrogates sex differences in T cell exhaustion.</p>
The development of extracellular vesicles (EV) for therapeutic applications is contingent upon the establishment of reproducible, scalable, and high-throughput methods for the production and purification of clinical grade EV. Methods including ultracentrifugation (U/C), ultrafiltration, immunoprecipitation, and size-exclusion chromatography (SEC) have been employed to isolate EV, each facing limitations such as efficiency, particle purity, lengthy processing time, and/or sample volume. We developed a cGMP-compatible method for the scalable production, concentration, and isolation of EV through a strategy involving bioreactor culture, tangential flow filtration (TFF), and preparative SEC. We applied this purification method for the isolation of engineered EV carrying multiple complexes of a novel human immunostimulatory cytokine-fusion protein, heterodimeric IL-15 (hetIL-15)/lactadherin. HEK293 cells stably expressing the fusion cytokine were cultured in a hollow-fibre bioreactor. Conditioned medium was collected and EV were isolated comparing three procedures: U/C, SEC, or TFF + SEC. SEC demonstrated comparable particle recovery, size distribution, and hetIL-15 density as U/C purification. Relative to U/C, SEC preparations achieved a 100-fold reduction in ferritin concentration, a major protein-complex contaminant. Comparative proteomics suggested that SEC additionally decreased the abundance of cytoplasmic proteins not associated with EV. Combination of TFF and SEC allowed for bulk processing of large starting volumes, and resulted in bioactive EV, without significant loss in particle yield or changes in size, morphology, and hetIL-15/lactadherin density. Taken together, the combination of bioreactor culture with TFF + SEC comprises a scalable, efficient method for the production of highly purified, bioactive EV carrying hetIL-15/lactadherin, which may be useful in targeted cancer immunotherapy approaches.
