Advances in mRNA therapeutics for cancer immunotherapy: From modification to delivery.

RNA vaccines have demonstrated their ability to solve the issues posed by the COVID-19 pandemic. This success has led to the renaissance of research into mRNA and their nanoformulations as potential therapeutic modalities for various diseases. The potential of mRNA as a template for synthesizing proteins and protein fragments for cancer immunotherapy is now being explored. Despite the promise, the use of mRNA in cancer immunotherapy is limited by challenges, such as low stability against extracellular RNases, poor delivery efficiency to the target organs and cells, short circulatory half-life, variable expression levels and duration. This review highlights recent advances in chemical modification and advanced delivery systems that are helping to address these challenges and unlock the biological and pharmacological potential of mRNA therapeutics in cancer immunotherapy. The review concludes by discussing future perspectives for mRNA-based cancer immunotherapy, which holds great promise as a next-generation therapeutic modality.

One-Pot Synthesis and Characterization of CuCrS2/ZnS Core/Shell Quantum Dots as New Blue-Emitting Sources.

In this paper, we introduce a new blue-emitting material, CuCrS2/ZnS QDs (CCS QDs). To obtain bright and stable photoluminescent probes, we prepared a core/shell structure; the synthesis was conducted in a one-pot system, using 1-dodecanethiol as a sulfur source and co-ligand. The CCS QDs exhibited a semi-spherical colloidal nanocrystalline shape with an average diameter of 9.0 nm and ZnS shell thickness of 1.6 nm. A maximum photoluminescence emission peak (PL max) was observed at 465 nm with an excitation wavelength of 400 nm and PLQY was 5% at an initial [Cr3+]/[Cu+] molar ratio of one in the core synthesis. With an off-stoichiometric modification for band gap engineering, the CCS QDs exhibited slightly blue-shifted PL emission spectra and PLQY was 10% with an increase in initial molar ratio of 2.0 (462 nm PL max). However, when the initial molar ratio exceeded two, the CCS QDs exhibited a lower photoluminescence quantum yield of 4.5% with 461 nm of PL max at the initial molar ratio of four due to the formation of non-emissive Cr2S3 nanoflakes.

Examining the temperature of embryo culture in in vitro fertilization: a randomized controlled trial comparing traditional core temperature (37°C) to a more physiologic, cooler temperature (36°C).

OBJECTIVE: To determine whether culture at a more physiologically cooler temperature, as suggested by limited human and animal data, would improve blastulation and pregnancy rates in human clinical IVF. DESIGN: Paired randomized controlled trial. SETTING: Academic. PATIENT(S): Infertile couples (n=52) with a female partner less than 42 years old with eight or more mature oocytes retrieved. INTERVENTION(S): Mature oocytes obtained from a single cohort of oocytes were randomly divided into two groups; one was cultured at 37°C and the other at 36°C from the time of ICSI to the time of embryo transfer or vitrification. Paired embryo transfers were accomplished by transferring one euploid embryo from each group. DNA fingerprinting was used as needed to determine the outcome for each embryo. MAIN OUTCOME MEASURE(S): Rate of development of expanded blastocysts suitable for transfer or vitrification (primary outcome), fertilization, aneuploidy, and sustained implantation. RESULT(S): A total of 805 mature oocytes were cultured; 399 at 36°C and 406 at 37°C. Paired analysis demonstrated a higher rate of usable blastocyst formation per zygote at 37°C (48.4%) vs. at 36°C (41.2%). Rates of fertilization, aneuploidy, and sustained implantation were equivalent. CONCLUSION(S): IVF culture at 36°C does not improve clinically relevant parameters of embryo development or sustained implantation rates. CLINICAL TRIAL REGISTRATION NUMBER: NCT01506089.

Spectral domain optical coherence tomography findings in bilateral peripheral cone dystrophy.

INTRODUCTION: To report spectral domain optical coherence (OCT) tomography findings in a case of bilateral peripheral cone dystrophy. CASE REPORT: A 21-year-old man complained of blurred visual fields on the lateral sides of both eyes. The patient's best-corrected visual acuity was 20/20 in both eyes. Fundus examination revealed mild temporal pallor, while fluorescein angiography did not show any abnormalities. Humphrey's visual field revealed a C-shaped scotoma. Photopic electroretinogram (ERG) and 30-Hz flicker revealed decreased cone function; however, scotopic ERG noted a normal response. Multifocal ERG revealed a relatively well-preserved macular area but with reduced amplitude in the peripheral areas of both eyes. On OCT, the photoreceptor inner and outer segment junction and retinal pigment epithelium (RPE) layers were well preserved in both eyes, except for a slight decrease in outer layer thickness. Moreover, on the macular thickness map obtained from Cirrus HD-OCT, a reduction in internal limiting membrane-RPE thickness that correlated well with visual field defects was revealed in both eyes. On the retinal nerve fiber layer (RNFL) thickness map obtained from Cirrus HD-OCT, the superior quadrant showed decreased RNFL thickness; however, thickness in the temporal quadrant corresponded with thickness in the macular area and was normal in both eyes. DISCUSSION: The OCT of a patient with peripheral cone dystrophy revealed decreased thickness of the macula with well-preserved retinal structures, which may be one of the distinctive features of this condition.

Analysis of spectral domain optical coherence tomography findings in occult macular dystrophy.

PURPOSE: To describe findings of spectral domain optical coherence tomography (SD-OCT) in the diagnosis of occult macular dystrophy (OMD). MATERIALS AND METHODS: Five visually disturbed patients who had shown normal findings in full field electroretinogram (ERG), fundus photography and fluorescein angiography but abnormal findings in multifocal ERG had undergone SD-OCT. The morphologic changes in retina and macular thickness were detected and imaged by SD-OCT. We analysed the results of SD-OCT including macular thickness, inner segment and outer segment (IS/OS) boundary and external limiting membrane (ELM). RESULTS: Mean age was 41.6 (22-63) in three male and two female patients in this study. Mean central foveal thickness was 142.9 (76-161) µm. All patients had shown decreased bowing of IS/OS boundary, and five of nine eyes were noted with disrupted IS/OS boundary of the photoreceptors according to SD-OCT. The interruption of external limiting membrane was noted in three of nine eyes. CONCLUSIONS: Spectral domain optical coherence tomography findings are useful in diagnosing OMD. Morphologic changes of the retina including deformity of the OS/IS boundary of the photoreceptors, disruption of the ELM and decreased foveal thickness may be important characteristics to consider when determining the pathophysiology and diagnosing criteria of OMD.

The suppressive effect of myeloid Elf-1-like factor (MEF) in osteogenic differentiation.

Myeloid Elf-1 like factor (MEF) is a member of the Ets transcription factor family. Ets family proteins control the expression of genes that are critical for biological processes such as proliferation, differentiation, and cell death. Some of Ets factors are also known to regulate bone development. In this study, we investigated the role of MEF in osteoblast differentiation. MEF expression was highest early in the differentiation of MC3T3-E1 osteoblasts and was reduced by treatment with BMP-2. The expression of MEF suppressed the alkaline phosphatase activity and expression induced by BMP-2 stimulation and mediated by Runx2. The expression of MEF also reduces osteocalcin mRNA levels, and mineralization in MC3T3-E1 cells. We found that the MEF-mediated suppression of osteogenic differentiation was critically related to Runx2 regulation. The MEF and Runx2 proteins physically interact to form a complex, and this interaction interferes with Runx2 binding to the cis-acting element OSE2 derived from the osteocalcin promoter. Co-transfection of MEF inhibited the 6xOSE2-luciferase reporter activity induced by Runx2. In addition, MEF stimulated the transcription of a negative mediator Msx2, and a transcriptional repressor, Mab21L1, and suppressed the transcription of a positive mediator, Dlx5 in osteoblast differentiation. MEF overexpression stimulated C2C12 cell proliferation. Together, our findings suggest that MEF promotes cell proliferation and functions as a negative regulator of osteogenic differentiation by directly interacting with Runx2 and suppressing its transcriptional activity.

Diverse pathways mediate chemotherapy-induced cell death in acute lymphoblastic leukemia cell lines.

Cancer cell resistance to chemotherapy may be mediated by defects in apoptotic pathways. A prior study showed that in vivo apoptosis of Acute Lymphoblastic Leukemia (ALL) blasts in response to chemotherapy could occur through diverse pathways including both p53-dependent and -independent mechanisms. In this study we investigated the apoptotic response in more detail by using a panel of ALL cell lines that differed in respect to p53 status. Upon exposure to a uniform stimulus, expression of apoptotic proteins, including the effector caspase-3, varied among ALL cell lines partly depending on p53 transcriptional activity and caspase-8 activation. Although the expression and contribution to apoptosis differed among known members of the apoptotic pathway, apoptosis was universally mediated by mitochondrial depolarization. The NFkappaB pathway was activated in response to chemotherapy but NFkappaB inhibition appeared to not influence chemosensitivity. This study further documents the highly variable nature of cell death programs in ALL and provides the foundation for cell death pathway modulation to improve ALL cure rates without increasing chemotherapy-related toxicity.

Biologic pathways associated with relapse in childhood acute lymphoblastic leukemia: a Children's Oncology Group study.

Outcome for children with childhood acute lymphoblastic leukemia (ALL) who relapse is poor. To gain insight into the mechanisms of relapse, we analyzed gene-expression profiles in 35 matched diagnosis/relapse pairs as well as 60 uniformly treated children at relapse using the Affymetrix platform. Matched-pair analyses revealed significant differences in the expression of genes involved in cell-cycle regulation, DNA repair, and apoptosis between diagnostic and early-relapse samples. Many of these pathways have been implicated in tumorigenesis previously and are attractive targets for intervention strategies. In contrast, no common pattern of changes was observed among late-relapse pairs. Early-relapse samples were more likely to be similar to their respective diagnostic sample while we noted greater divergence in gene-expression patterns among late-relapse pairs. Comparison of expression profiles of early- versus late-relapse samples indicated that early-relapse clones were characterized by overexpression of biologic pathways associated with cell-cycle regulation. These results suggest that early-relapse results from the emergence of a related clone, characterized by the up-regulation of genes mediating cell proliferation. In contrast, late relapse appears to be mediated by diverse pathways.

Glutamate-induced glutamine synthetase expression in retinal Muller cells after short-term ocular hypertension in the rat.

PURPOSE: To determine the effect of intraocular pressure (IOP) elevation on glutamate-induced expression of glutamine synthetase (GS) in retinal Müller cells of rat eyes. METHODS: Six groups of three rats each were studied. Group I was a normal control group. In Group II, one eye received an intravitreal glutamate injection (75 nmoles) while the contralateral eye served as control. In Groups III and IV, IOP was raised in one eye by episcleral vein cauterization. Moderately elevated IOP was maintained for 1 day in Group III or 1 week in Group IV (35 +/- 1.9-45 +/- 5.2 mm Hg). An additional two groups of rats received bilateral intravitreal glutamate injections (75 nmoles) immediately (Group V), or 6 days (Group VI), after induction of IOP elevation in one eye. One day after glutamate injection the rats in all groups were killed, and the eyes enucleated and fixed. Retinas from left and right eyes of each animal were embedded together in LR White resin (Ted Pella, Redding, CA). Sections were processed for GS immunolabeling with antibodies to GS by two-stage immunogold labeling with silver enhancement. Images of labeled retinas from the two eyes were captured under identical light microscopic conditions and the GS immunoreactivity in Müller cells was compared between the left and right retinas in the same section by image analysis. An additional five rats were included in Group II and the retinas were analyzed by Western blot analysis to confirm immunohistochemical findings. RESULTS: Similar to the finding in the control group (Group I), the GS immunoreactivity of the left and right eyes of Group III and IV remained unchanged even though the right eyes in the two groups had elevation of IOP lasting for 24 hours and 1 week, respectively. However, GS levels were significantly increased by 40 +/- 5.7% in normotensive eyes 24 hours after intravitreal injection of glutamate (Group II). The rise in GS immunoreactivity was abolished in eyes with acute IOP elevation (Group V). In contrast, when the eyes were exposed to high IOP for 1 week (Group VI), the glutamate-induced increase in GS immunoreactivity was restored. CONCLUSIONS: Elevated levels of vitreal glutamate can increase the expression of GS in retinal Müller cells. This increase was blocked if IOP was acutely elevated for 24 hours but was restored if IOP remained elevated for 1 week. This finding suggests that moderate elevation of IOP causes only short-term functional changes of glutamate metabolism (amidation) by retinal Müller cells. However, it is not known to what extent endogenous extracellular glutamate can regulate GS expression in normal eyes or in eyes with glaucoma.