Combinatorial inhibition of BTK, PI3K-AKT and BRD4-MYC as a strategy for treatment of mantle cell lymphoma

Mantle cell lymphoma (MCL) is a subtype of non-Hodgkin’s lymphoma characterized by poor prognosis. The complexity of MCL pathogenesis arises from aberrant activities of diverse signaling pathways, including BTK, PI3K–AKT–mTOR and MYC-BRD4. Here, we report that MCL-related signaling pathways can be altered by a single small molecule inhibitor, SRX3305. Binding and kinase activities along with resonance changes in NMR experiments reveal that SRX3305 targets both bromodomains of BRD4 and is highly potent in inhibition of the PI3K isoforms α, γ and δ, as well as BTK and the drug-resistant BTK mutant. Preclinical investigations herein reveal that SRX3305 perturbs the cell cycle, promotes apoptosis in MCL cell lines and shows dose dependent anti-proliferative activity in both MCL and drug-resistant MCL cells. Our findings underscore the effectiveness of novel multi-action small molecule inhibitors for potential treatment of MCL.

Optimization of miR-22 expression cassette for rAAV delivery on diabetes

MicroRNA-22 (miR-22) was suggested to be important for type 2 diabetes but its functions for this disease remained unclear. Recombinant adeno-associated virus (rAAV)-mediated miR delivery is a powerful approach to study miR functions in vivo, however, the overexpression of miR-22 by rAAV remains challenging because it is one of the most abundant miRs in the liver. In this study, a series of expression cassettes were designed and compared. It was shown that different lengths of primary miR-22 were overexpressed in HEK293 and HeLa cells but the longer ones were more efficiently expressed. miR-22 may be placed in either introns or the 3′ UTR of a transgene for efficient overexpression. RNA polymerase III or II promoters were successfully utilized for miR expression but the latter showed higher expression levels in cell lines. Specifically, miR-22 was expressed efficiently together with an EGFP gene. After screening, a liver-specific TTR promoter was chosen to overexpress miR-22 in diabetic mice fed a high-fat diet. It was shown that miR-22 was overexpressed 2-3 folds which improved the insulin sensitivity significantly. The approach utilized in this study to optimize miR overexpression is a powerful tool for the creation of efficient rAAV vectors for the other miRs.

Cholesterol modified DP7 and pantothenic acid induce dendritic cell homing to enhance the efficacy of dendritic cell vaccines

Dendritic cell (DC)-based cancer vaccines have so far achieved good therapeutic effects in animal experiments and early clinical trials for certain malignant tumors. However, the overall objective response rate in clinical trials rarely exceeds 15%. The poor efficiency of DC migration to lymph nodes (LNs) (< 5%) is one of the main factors limiting the effectiveness of DC vaccines. Therefore, increasing the efficiency of DC migration is expected to further enhance the efficacy of DC vaccines. Here, we used DP7-C (cholesterol modified VQWRIRVAVIRK), which can promote DC migration, as a medium. Through multiomics sequencing and biological experiments, we found that it is the metabolite pantothenic acid (PA) that improves the migration and effectiveness of DC vaccines. We clarified that both DP7-C and PA regulate DC migration by regulating the chemokine receptor CXCR2 and inhibiting miR-142a-3p to affect the NF-κB signaling pathway. This study will lay the foundation for the subsequent use of DP7-C as a universal substance to promote DC migration, further enhance the antitumor effect of DC vaccines, and solve the bottleneck problem of the low migration efficiency and unsatisfactory clinical response rate of DC vaccines.

Untangling the KRAS mutated lung cancer subsets and its therapeutic implications

The Kirsten rat sarcoma virus transforming protein (KRAS) mutations (predominate in codons 12, 13, and 61) and genomically drive nearly one-third of lung carcinomas. These mutations have complex functions in tumorigenesis, and influence the tumor response to chemotherapy and tyrosine kinase inhibitors resulting in a poorer patient prognosis. Recent attempts using targeted therapies against KRAS alone have met with little success. The existence of specific subsets of lung cancer based on KRAS mutations and coexisting mutations are suggested. Their interactions need further elaboration before newer promising targeted therapies for KRAS mutant lung cancers can be used as earlier lines of therapy. We summarize the existing knowledge of KRAS mutations and their coexisting mutations that is relevant to lung cancer treatment, in this review. We elaborate on the prognostic impact of clinical and pathologic characteristics of lung cancer patients associated with KRAS mutations. We briefly review the currently available techniques for KRAS mutation detection on biopsy and cytology samples. Finally, we discuss the new therapeutic strategies for targeting KRAS-mutant non-small cell lung cancer (NSCLC). These may herald a new era in the treatment of KRASG12Cmutated NSCLC as well as be helpful to develop demographic subsets to predict targeted therapies and prognosis of lung cancer patients.

TIPE polarity proteins are required for mucosal deployment of T lymphocytes and mucosal defense against bacterial infection

Mucosal surfaces are continuously exposed to, and challenged by, numerous commensal and pathogenic organisms. To guard against infections, a majority of the thymus-derived T lymphocytes are deployed at the mucosa. Although chemokines are known to be involved in the mucosal lymphocyte deployment, it is not clear whether lymphocytes enter the mucosa through directed migration or enhanced random migration. Here we report that TIPE (tumor necrosis factor-α-induced protein 8 (TNFAIP8)-like) proteins mediate directed migration of T lymphocytes into lung mucosa, and they are crucial for mucosal immune defense against Streptococcus pneumoniae infection. Knockout of both Tnfaip8 and Tipe2, which encode polarity proteins that control the directionality of lymphocyte migration, significantly reduced the numbers of T lymphocytes in the lung of mice. Compared with wild-type mice, Tnfaip8−/−Tipe2−/− mice also developed more severe infection with more pathogens entering blood circulation upon nasal Streptococcus pneumoniae challenge. Single-cell RNA-sequencing analysis revealed that TIPE proteins selectively affected mucosal homing of a unique subpopulation of T cells, called “T cells-2”, which expressed high levels of Ccr9, Tcf7, and Rag1/2 genes. TNFAIP8 and TIPE2 appeared to have overlapping functions since deficiency in both yielded the strongest phenotype. These data demonstrate that TIPE family of proteins are crucial for lung mucosal immunity. Strategies targeting TIPE proteins may help develop mucosal vaccines or treat inflammatory diseases of the lung.

Roles of host mitochondria in the development of COVID-19 pathology: Could mitochondria be a potential therapeutic target?

The recent emergence of severe acute respiratory syndrome-Corona Virus 2 (SARS-CoV-2) in late 2019 and its spread worldwide caused an acute pandemic of Coronavirus disease 19 (COVID-19). Since then, COVID-19 has been under intense scrutiny as its outbreak led to significant changes in healthcare, social activities, and economic settings worldwide. Although angiotensin-converting enzyme-2 (ACE-2) receptor is shown to be the primary port of SARS-CoV-2 entry in cells, the mechanisms behind the establishment and pathologies of COVID-19 are poorly understood. As recent studies have shown that host mitochondria play an essential role in virus-mediated innate immune response, pathologies, and infection, in this review, we will discuss in detail the entry and progression of SARS-CoV-2 and how mitochondria could play roles in COVID-19 disease. We will also review the potential interactions between SARS-CoV-2 and mitochondria and discuss possible treatments, including whether mitochondria as a potential therapeutic target in COVID-19. Understanding SARS-CoV-2 and mitochondrial interactions mediated virus establishment, inflammation, and other consequences may provide a unique mechanism and conceptual advancement in finding a novel treatment for COVID-19.

Random-PE: an efficient integration of random sequences into mammalian genome by prime editing

Prime editing (PE) enables efficiently targeted introduction of multiple types of small-sized genetic change without requiring double-strand breaks or donor templates. Here we designed a simple strategy to introduce random DNA sequences into targeted genomic loci by prime editing, which we named random prime editing (Random-PE). In our strategy, the prime editing guide RNA (pegRNA) was engineered to harbor random sequences between the primer binding sequence (PBS) and homologous arm (HA) of the reverse transcriptase templates. With these pegRNAs, we achieved efficient targeted insertion or substitution of random sequences with different lengths, ranging from 5 to 10, in mammalian cells. Importantly, the diversity of inserted sequences is well preserved. By fine-tuning the design of random sequences, we were able to make simultaneously insertions or substitutions of random sequences in multiple sites, allowing in situ evolution of multiple positions in a given protein. Therefore, these results provide a framework for targeted integration of random sequences into genomes, which can be redirected for manifold applications, such as in situ protospacer adjacent motif (PAM) library construction, enhancer screening, and DNA barcoding.

Natural variant frequencies across domains from different sarcomere proteins cross-correlate to identify inter-protein contacts associated with cardiac muscle function and disease

Coordinated sarcomere proteins produce contraction force for muscle shortening. In human ventriculum they include the cardiac myosin motor (βmys), repetitively converting ATP free energy into work, and myosin binding protein C (MYBPC3) that in complex with βmys is regulatory. Single nucleotide variants (SNVs) causing hereditary heart diseases frequently target this protein pair. The βmys/MYBPC3 complex models a regulated motor and is used here to study how the proteins couple. SNVs in βmys or MYBPC3 survey human populations worldwide. Their protein expression modifies domain structure affecting phenotype and pathogenicity outcomes. When the SNV modified domain locates to inter-protein contacts it could affect complex coordination. Domains involved, one in βmys the other in MYBPC3, form coordinated domains (co-domains). Co-domain bilateral structure implies the possibility for a shared impact from SNV modification in either domain suggesting a correlated response to a common perturbation could identify their location. Genetic divergence over human populations is proposed to perturb SNV probability coupling that is detected by cross-correlation in 2D correlation genetics (2D-CG). SNV probability data and 2D-CG identify three critical sites, two in MYBPC3 with links to several domains across the βmys motor, and, one in βmys with links to the MYBPC3 regulatory domain. MYBPC3 sites are hinges sterically enabling regulatory interactions with βmys. The βmys site is the actin binding C-loop (residues 359-377). The C-loop is a trigger for actin-activated myosin ATPase and a contraction velocity modulator. Co-domain identification implies their spatial proximity suggesting a novel approach for in vivo protein complex structure determination.

Targeting cardiomyocyte proliferation as a key approach of promoting heart repair after injury

Cardiovascular diseases such as myocardial infarction (MI) is a major contributor to human mortality and morbidity. The mammalian adult heart almost loses its plasticity to appreciably regenerate new cardiomyocytes after injuries, such as MI and heart failure. The neonatal heart exhibits robust proliferative capacity when exposed to varying forms of myocardial damage. The ability of the neonatal heart to repair the injury and prevent pathological left ventricular remodeling leads to preserved or improved cardiac function. Therefore, promoting cardiomyocyte proliferation after injuries to reinitiate the process of cardiomyocyte regeneration, and suppress heart failure and other serious cardiovascular problems have become the primary goal of many researchers. Here, we review recent studies in this field and summarize the factors that act upon the proliferation of cardiomyocytes and cardiac repair after injury and discuss the new possibilities for potential clinical treatment strategies for cardiovascular diseases.