CD33‐targeting extracellular vesicles deliver antisense oligonucleotides against FLT3‐ITD and miR‐125b for specific treatment of acute myeloid leukaemia

IntroductionAcute Myeloid Leukaemia (AML) is the most common blood cancer in adults. Although 2 out of 3 AML patients go into total remission after chemotherapies and targeted therapies, the disease recurs in 60%–65% of younger adult patients within 3 years after diagnosis with a dramatically decreased survival rate. Therapeutic oligonucleotides are promising treatments under development for AML as they can be designed to silence oncogenes with high specificity and flexibility. However, there are not many well validated approaches for safely and efficiently delivering oligonucleotide drugs. This issue could be resolved by utilizing a new generation of delivery vehicles such as extracellular vesicles (EVs).MethodsIn this study, we harness red blood cell‐derived EVs (RBCEVs) and engineer them via exogenous drug loading and surface functionalization to develop an efficient drug delivery system for AML. Particularly, EVs are designed to target CD33, a common surface marker with elevated expression in AML cells via the conjugation of a CD33‐binding monoclonal antibody onto the EV surface.ResultsThe conjugation of RBCEVs with the CD33‐binding antibody significantly increases the uptake of RBCEVs by CD33‐positive AML cells, but not by CD33‐negative cells. We also load CD33‐targeting RBCEVs with antisense oligonucleotides (ASOs) targeting FLT3‐ITD or miR‐125b, 2 common oncogenes in AML, and demonstrate that the engineered EVs improve leukaemia suppression in in vitro and in vivo models of AML.ConclusionTargeted RBCEVs represent an innovative, efficient, and versatile delivery platform for therapeutic ASOs and can expedite the clinical translation of oligonucleotide drugs for AML treatments by overcoming current obstacles in oligonucleotide delivery.

In this study, we harness red blood cell‐derived EVs (RBCEVs) and engineer them with surface functionalization and exogenous drug loading to develop an efficient drug delivery system for AML. Anti‐CD33 antibody was conjugated to RBCEVs using an enzymatic method combined with the streptavidin‐biotin system. We load the antibody conjugated RBCEVs with ASOs targeting FLT3‐ITD or miR‐125b, 2 common oncogenes in AML, and demonstrate that the treatment with engineered EVs improve leukaemia suppression both in vitro and in vivo.     

Virtual assessment,training, and evaluation of clinical laboratory technologists amidst peak Covid-19 pandemic: An observational study

Medical technologists are considered a neglected group when it comes to academic interventions. We developed and implemented an educational intervention and assessment for the technologists based on an online questionnaire as a pre-test consisting of questions related to knowledge (n=5), attitude (n=3), and practices (n=4) of daily internal quality control (QC) monitoring via Google Docs survey tool. This study served multiple purposes. It allowed keeping the technologists engaged during the peak of the COVID-19 pandemic while also improving the knowledge, attitude, and practices about the internal quality control using Bio-Rad Unity Real Time (URT) QC software. Subjects were graded based on the scores they received out of 100 (0-60 = poor; 61-79 = good; 80-100 = excellent). Training materials, i.e., a set of 5 videos every week via e-mail, were circulated. A voice-over PowerPoint presentation was also shared for easy comprehension. This activity was repeated after one month. A post-test was administered to assess the improvement. The study results show significant improvement in the technologists'' performance after the intervention.     

Shrinkage of the non-malignant prostate gland volume after receiving incidental radiotherapy for rectal cancer

BackgroundThe purpose of this study was to assess the impact of coincidental radiotherapy on the volume of the non-malignant prostate gland in rectal cancer patients treated with neo-adjuvant radiotherapy.Materials and methodsIn this retrospective analysis, thirty male patients with rectal cancer who had neoadjuvant radiotherapy met the inclusion criteria. These patients had pre-treatment magnetic resonance imaging (MRI) and at least one post-treatment MRI of the pelvis and the whole of their prostate volume received the full prescribed radiotherapy dose; 45 Gy in 25 fractions (n = 22), 45 Gy in 20 fractions (n = 4) and 25 Gy in 5 fractions (n = 4).ResultsThe median age of this patient cohort was 66 years (range: 30–87). With a median interval between pre-treatment MRI and first MRI post-treatment of 2 months (range: 1–11), the mean prostate volume reduced from 36.1 cm³ [standard deviation (SD) 14.2] pre-radiotherapy to 31.3 cm³ (SD 13.0) post radiotherapy and this difference was significant (p = 0.0004).ConclusionRadiotherapy may cause shrinkage in volume of normal (non-malignant) prostate. Further research is required in this field, since these results may be of some comfort to men contemplating the consequences of radiotherapy on their quality of life. The authors suggest recording flow-rate and international prostate symptom score (IPSS) during rectal radiotherapy as a next step.     

Medium optimization for chitinase production from Trichoderma virens using central composite design

Medium development for chitinase production by Trichoderma virens was first carried out using conventional method of one-factor-at-a-time. The medium was further optimized using Central Composite Design in which response surface was generated later from the derived model. An experimental design of four variables including various initial pH values, chitin, ammonium sulphate, and methanol concentrations were created using Design Expert® Software, Version 6.0. The design consists of 30 experiments, which include 6 replicates at center points. The optimal value for each variable are 3.0 g/L, chitin; 0.1 g/L, ammonium sulphate; 0.4% (v/v), methanol; and initial pH, 4.0 with predicted chitinase activity of 0.1495 U/mL. These predicted parameters were tested in the laboratory and the final chitinase activity obtained was 0.1471 U/mL, which is almost reaching the predicted value. The optimal medium design showed an improvement of chitinase activity of 80.9% compared to activity obtained from the original Absidia medium composition.     

Multi-point enzyme immobilization,surface chemistry,and novel platforms: a paradigm shift in biocatalyst design

Engineering enzymes with improved catalytic properties in non-natural environments have been concerned with their diverse industrial and biotechnological applications. Immobilization represents a promising but straightforward route, and immobilized biocatalysts often display higher activities and stabilities compared to free enzymes. Owing to their unique physicochemical characteristics, including the high-specific surface area, exceptional chemical, electrical, and mechanical properties, efficient enzyme loading, and multivalent functionalization, nano-based materials are postulated as suitable carriers for biomolecules or enzyme immobilization. Enzymes immobilized on nanomaterial-based supports are more robust, stable, and recoverable than their pristine counterparts, and are even used for continuous catalytic processes. Furthermore, the unique intrinsic properties of nanomaterials, particularly nanoparticles, also confer the immobilized enzymes to be used for their broader applications. Herein, an effort has been made to present novel potentialities of multi-point enzyme immobilization in the current biotechnological sector. Various nano-based platforms for enzyme/biomolecule immobilization are discussed in the second part of the review. In summary, recent developments in the use of nanomaterials as new carriers to construct robust nano-biocatalytic systems are reviewed, and future trends are pointed out in this article.     

Seed Treatment with Auxins Modulates Growth and Ion Partitioning in Salt-stressed Wheat Plants

Experiments were performed to determine whether seed priming with different concentrations (100, 150, and 200 mg/L) ofauxins (indoleacetic acid (IAA), indolebutyric acid (IBA), or their precursor tryptophane (Trp)) could alter salinity inducedperturbances in salicylic acid and ion concentrations and, hence, growth in wheat (Triticum aestivum L.) cultivars, namelyM. H.-97 (salt intolerant) and Inqlab-91 (salt tolerant). Primed and non-primed seeds were sown in Petri dishes in a growthroom, as well as in a field treated with 15 dS/m NaCl salinity. All priming agents, except IBA, increased the final germinationpercentage in both cultivars. The seedlings of either cultivar raised from Trp-treated seeds had greater dry biomass whenunder salt stress. In field experiments, Trp priming was much more effective in mediating the increase in grain yield,irrespective of the cultivar, under salt stress. The alleviatory effect of Trp was found to be associated with reduced uptakeof Na~ in the roots and subsequent translocation to the shoots, as well as increased partitioning of Ca~(2 )in the roots ofsalt-stressed wheat plants. Plants of both cultivars raised from Trp-and IAA-treated seeds accumulated free salicylic acidin their leaves when under salt stress. Overall, the Trp priming-induced improvement in germination and the subsequentgrowth of wheat plants could be related to ion homeostasis when under salt stress. The possible involvement of salicylicacid in the Trp priming-induced better growth under conditions of salt stress is discussed.     

Effects of three different cultivars of cruciferous plants on the age‐stage,two‐sex life table traits of Plutella xylostella (L.) (Lepidoptera: Plutellidae)

Plutella xylostella is an important pest of cruciferous crops worldwide. However, information regarding the age‐stage, two‐sex life parameters of P. xylostella, which is vital for designing more effective control methods, is currently lacking. The present study reports age‐stage, two‐sex life table parameters for P. xylostella on napa cabbage (Brassica oleracea var. napa), white cabbage (B. oleracea var. capitata), and cauliflower (B. oleracea var. botrytis) under laboratory conditions at 25 ± 2°C, 50–60% relative humidity, and a 16‐h light : 8‐h dark photoperiod. The time for development from an egg to a male or female adult P. xylostella on white cabbage (mean [± SE] 41.15 ± 0.54 and 39.50 ± 0.54 days, respectively) was significantly longer than that on cauliflower and napa cabbage. Furthermore, P. xylostella fecundity on cauliflower (261.90 ± 4.53 eggs female) was significantly highest than on napa cabbage and white cabbage. Intrinsic rate of increase (r) and finite rate of increase (λ) were highest on cauliflower 0.182 day^?1 and 1.199 day^?1 respectively as comparison to napa cabbage and white cabbage. The highest gross reproductive rate (GRR) and net reproductive rates (R₀) of P. xylostella 65.87 and 52.58 respectively on cauliflower then those of other hosts. The findings of the present study indicate that cauliflower is the most suitable cultivar (host) for the development of P. xylostella. Based on these findings, crops like cauliflower can be used as trap crops when napa cabbage and white cabbage are the main crops.