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Nature has been always a great source of possible lead compounds to develop new drugs against several diseases. Here we report the identification of a natural compound, membranoid G, derived from the Antarctic sponge Dendrilla antarctica displaying an in vitro inhibitory activity against human DNA topo-isomerase 1B. The experiments indicate that membranoid G, when pre-incubated with the enzyme, strongly and irreversibly inhibits the relaxation of supercoiled DNA. This compound completely inhibits the cleavage step of the enzyme catalytic mechanism by preventing protein binding to the DNA. Membranoid G displays also a cytotoxic effect on tumour cell lines, suggesting its use as a possible lead compound to develop new anticancer drugs. [ABSTRACT FROM AUTHOR]

KRAS mutations hinder therapeutic efficacy of epidermal growth factor receptor (EGFR)‐specific monoclonal antibodies cetuximab and panitumumab‐based immunotherapy of EGFR+ cancers. Although cetuximab inhibits KRAS‐mutated cancer cell growth in vitro by natural killer (NK) cell‐mediated antibody‐dependent cellular cytotoxicity (ADCC), KRAS‐mutated colorectal carcinoma (CRC) cells escape NK cell immunosurveillance in vivo. To overcome this limitation, we used cetuximab and panitumumab to redirect Fcγ chimeric receptor (CR) T cells against KRAS‐mutated HCT116 colorectal cancer (CRC) cells. We compared four polymorphic Fcγ‐CR constructs including CD16158F‐CR, CD16158V‐CR, CD32131H‐CR, and CD32131R‐CR transduced into T cells by retroviral vectors. Percentages of transduced T cells expressing CD32131H‐CR (83.5 ± 9.5) and CD32131R‐CR (77.7 ± 13.2) were significantly higher than those expressing with CD16158F‐CR (30.3 ± 10.2) and CD16158V‐CR (51.7 ± 13.7) (p < 0.003). CD32131R‐CR T cells specifically bound soluble cetuximab and panitumumab. However, only CD16158V‐CR T cells released high levels of interferon gamma (IFNγ = 1,145.5 pg/ml ±16.5 pg/ml, p < 0.001) and tumor necrosis factor alpha (TNFα = 614 pg/ml ± 21 pg/ml, p < 0.001) upon incubation with cetuximab‐opsonized HCT116 cells. Moreover, only CD16158V‐CR T cells combined with cetuximab killed HCT116 cells and A549 KRAS‐mutated cells in vitro. CD16158V‐CR T cells also effectively controlled subcutaneous growth of HCT116 cells in CB17‐SCID mice in vivo. Thus, CD16158V‐CR T cells combined with cetuximab represent useful reagents to develop innovative EGFR+KRAS‐mutated CRC immunotherapies. What's new? KRAS mutations downstream the epidermal growth factor receptor (EGFR) represent the major limitation in monoclonal antibody‐targeted therapy of EGFR+ colorectal cancers. To restore the sensitivity of KRAS‐mutated cancer cells to EGFR‐specific mAbs, here the authors explore different strategies based on the generation of extracellular CD16‐chimeric receptors linked to intracellular signalling and subsequent transduction into T cells. They demonstrate that T cells engineered with CD16158V‐chimeric receptor can be redirected by EGFR‐specific mAbs toward EGFR+ cancer cells, overcoming the limitation conferred by KRAS mutations in vitro and in vivo. This study might help develop innovative and effective treatments of EGFR+ KRAS‐mutated cancers. [ABSTRACT FROM AUTHOR]

Cetuximab and panitumumab bind the human epidermal growth factor receptor (EGFR). Although the chimeric cetuximab (IgG1) triggers antibody‐dependent‐cellular‐cytotoxicity (ADCC) of EGFR positive target cells, panitumumab (a human IgG2) does not. The inability of panitumumab to trigger ADCC reflects the poor binding affinity of human IgG2 Fc for the FcγRIII (CD16) on natural killer (NK) cells. However, both human IgG1 and IgG2 bind the FcγRII (CD32A) to a similar extent. Our study compares the ability of T cells, engineered with a novel low‐affinity CD32A131R‐chimeric receptor (CR), and those engineered with the low‐affinity CD16158F‐CR T cells, in eliminating EGFR positive epithelial cancer cells (ECCs) in combination with cetuximab or panitumumab. After T‐cell transduction, the percentage of CD32A131R‐CR T cells was 74 ± 10%, whereas the percentage of CD16158F‐CR T cells was 46 ± 15%. Only CD32A131R‐CR T cells bound panitumumab. CD32A131R‐CR T cells combined with the mAb 8.26 (anti‐CD32) and CD16158F‐CR T cells combined with the mAb 3g8 (anti‐CD16) eliminated colorectal carcinoma (CRC), HCT116FcγR+ cells, in a reverse ADCC assay in vitro. Crosslinking of CD32A131R‐CR on T cells by cetuximab or panitumumab and CD16158F‐CR T cells by cetuximab induced elimination of triple negative breast cancer (TNBC) MDA‐MB‐468 cells, and the secretion of interferon gamma and tumor necrosis factor alpha. Neither cetuximab nor panitumumab induced Fcγ‐CR T antitumor activity against Kirsten rat sarcoma (KRAS)‐mutated HCT116, nonsmall‐cell‐lung‐cancer, A549 and TNBC, MDA‐MB‐231 cells. The ADCC of Fcγ‐CR T cells was associated with the overexpression of EGFR on ECCs. In conclusion, CD32A131R‐CR T cells are efficiently redirected by cetuximab or panitumumab against breast cancer cells overexpressing EGFR. What's new? The role of antibody‐dependent cellular cytotoxicity in the antitumor activity of tumor antigen‐specific monoclonal antibodies has stimulated interest in genetically engineering T cells with the CD16 chimeric receptor (CD16‐CR). Here, the authors expand the application of this methodology to triple negative breast cancer (TNBC) overexpressing EGFR by utilizing a novel CD32A131R‐CR in combination with anti‐EGFR mAbs. The study supports the use of CD32A131R‐CR T cells combined with either panitumumab or cetuximab—an IgG2 and IgG1 monoclonal antibody, respectively—for targeting TNBC cells overexpressing EGFR. The results may be utilized as a platform for designing therapies targeting TNBC overexpressing EGFR. [ABSTRACT FROM AUTHOR]

DNA topoisomerase I enzymes relieve the torsional strain in DNA; they are essential for fundamental molecular processes such as DNA replication, transcription, recombination, and chromosome condensation; and act by cleaving and then religating DNA strands. Over the past few decades, scientists have focused on the DNA topoisomerases biological functions and established a unique role of Type I DNA topoisomerases in regulating gene expression and DNA chromosome condensation. Moreover, the human enzyme is being investigated as a target for cancer chemotherapy. The active site tyrosine is responsible for initiating two transesterification reactions to cleave and then religate the DNA backbone, allowing the release of superhelical tension. The different steps of the catalytic mechanism are affected by various inhibitors; some of them prevent the interaction between the enzyme and the DNA while others act as poisons, leading to TopI-DNA lesions, breakage of DNA, and eventually cellular death. In this review, our goal is to provide an overview of mechanism of human topoisomerase IB action together with the different types of inhibitors and their effect on the enzyme functionality., Competing Interests: All authors declared that there are no conflicts of interest., (© The Author(s) 2020.)

Background: DNA topoisomerases 1B are a class of ubiquitous enzyme that solves the topological problems associated with biological processes such as replication, transcription and recombination. Numerous sequence alignment of topoisomerase 1B from different species shows that the lengths of different domains as well as their amino acids sequences are quite different. In the present study a hybrid enzyme, generated by swapping the N-terminal of Plasmodium falciparum into the corresponding domain of the human, has been characterized., Methods: The chimeric enzyme was generated using different sets of PCR. The in vitro characterization was carried out using different DNA substrate including radio-labelled oligonucleotides., Results: The chimeric enzyme displayed slower relaxation activity, cleavage and re-ligation kinetics strongly perturbed when compared to the human enzyme., Conclusion: These results indicate that the N-terminal domain has a crucial role in modulating topoisomerase activity in different species.