In this issue:- Knock-in mouse feature
- Latest publications
- Ozgene PhD graduate
- Timeline update
Knock-in mice to the rescue
Knock-in mice have an insertion in a specific locus in order to produce genetically modified mice for various applications in medical research. Many knock-in mouse models are utilised to study and develop treatments for human diseases and we would like to introduce two that have been featured in recent publications.
Neuroblastoma knock-in mouse
Neuroblastoma is a solid tumour childhood cancer which accounts for 8–10% of all childhood cancer deaths. It is the most commonly diagnosed cancer in infants under one year, with an average age of diagnosis at two years. Neuroblastoma develops in the sympathetic nervous system and often starts in the adrenal gland, subsequently spreading to tissues in the abdomen, chest, pelvis, and neck. Low-risk neuroblastoma typically has a good outcome with surgery or simply observation. In high-risk neuroblastoma, however, chances of survival are less than 40% despite treatment. There are no known reasons or clear environmental links for neuroblastoma. In some cases, it runs in families due to mutations in anaplastic lymphoma kinase (ALK) or paired like homeobox 2B (PHOX2B) genes.
Dr. Ruth Palmer is focused on understanding the importance and function of ALK signalling during development and disease. Dr. Palmer is a Professor at the Gothenburg University in Sweden and her Lab at the Sahlgrenska Academy is primarily interested in intercellular signal transduction events mediated by the ALK receptor.
In a recent paper published in the EMBO Journal, the Palmer Lab together with the Hallberg and Van den Eynden labs investigated the role of ALK and LTK ligand 2 (ALKAL2) in the progression of neuroblastoma utilising a knock-in mouse model designed to constitutively express ALKAL2, developed by Ozgene. ALK has been identified as a neuroblastoma oncogene, which has raised the possibility of using ALK tyrosine kinase inhibitors (TKIs) in the treatment of patients with activating ALK mutations. 8-10% of neuroblastoma patients are ALK-positive, and the figure increases in the relapsed population. ALK is activated by ALKAL2 located on chromosome 2p, in the "2p-gain" region associated with neuroblastoma.
Dysregulation of ALKAL2 in neuroblastoma has not been previously investigated. Therefore, Prof. Palmer and her collaborators tested whether upregulated ALKAL2 would increase neuroblastoma progression in the absence of ALK mutation. The study shows that ALKAL2 overexpression in knock-in mice does indeed drive ALK TKI-sensitive neuroblastoma even without ALK mutation. The results suggest that neuroblastoma patients that do not have ALK mutation, such as those exhibiting 2p-gain, may also benefit from ALK TKI-based therapeutic treatment, giving hope to more families affected by neuroblastoma.
For more information on Prof. Palmer’s research, please see the publication below and visit the Ruth Palmer Lab website. For information on knock-in mouse models, visit Ozgene's knock-in mouse services.
Knock-in mouse for kidney study
The kidney plays an incredible role in maintaining a steady state of internal, physical-chemical conditions in our living system. The renal distal convoluted tubule (DCT), a part of the nephron structure in kidneys is critical for the fine-tuning of urinary ion excretion and arterial blood pressure regulation. Alas, we are only scratching the surface of understanding the regulatory process that governs the DCT function, especially regarding the transcriptional control mechanisms.
Dr. Johannes Loffing, Professor at the Institute of Anatomy at the University of Zürich, has dedicated his career to understanding ion balance in the kidneys and how deranged underlying mechanisms lead to human diseases, like arterial hypertension. Dr. Loffing is also a Director of the NCCR Kidney.CH.
One of his many published research works shows that the transcription factor Prox-1 gene is highly expressed in the DCT of adult mice. Prox-1 is a transcription factor that is essential in developmental processes in many mammalian organs. This sparked the idea to investigate if the highly expressed gene may contribute to the transcriptional regulation of the DCT function and structure, so Dr. Loffing and his team partnered with Ozgene to develop a knock-in mouse model that allowed an inducible, DCT-specific deletion of Prox-1 in the adult kidney.
The results indicate the deletion of Prox-1 has no obvious impact on the DCT structure and growth independent in both newborn and adult mice. However, Prox-1 deficient mice display significant hypomagnesemia with profound downregulation at both mRNA and protein levels of the DCT-specific apical Mg2+ channel TRPM6 and the NaCl cotransporter (NCC).
This discovery will give insights into genetic and acquired diseases affecting the DCT, and sets an important foundation for future studies on other DCT-enriched transcription factors in regulatory pathways governing kidney function. For more information on Dr. Loffing and his team research, please see publication below or visit the Institute of Anatomy website .
For information on knock-in mouse models, visit Ozgene's knock-in mouse services.
EMBO J. 2021 Jan 7. - FEATURED
ALK ligand ALKAL2 potentiates MYCN-driven neuroblastoma in the absence of ALK mutation. Borenäs M, Umapathy G, Lai WY, Lind DE, Witek B, Guan J, Mendoza-Garcia P, Masudi T, Claeys A, Chuang TP, Wakil A El, Arefin B, Fransson S, Koster J, Johansson M, Gaarder J, den Eynden J Van, Hallberg B, Palmer RH. - Sweden, China, Belgium, The Netherlands. [read]
Pflugers Arch. 2020 Nov 16. - FEATURED
Deletion of the transcription factor Prox-1 specifically in the renal distal convoluted tubule causes hypomagnesemia via reduced expression of TRPM6 and NCC. Schnoz C, Moser S, Kratschmar DV, Odermatt A, Loffing-Cueni D, Loffing J. - Switzerland. [read]
PLoS One. 2020 Dec 31.
Intense light as anticoagulant therapy in humans. Oyama Y, Shuff S, Davizon-Castillo P, Clendenen N, Eckle T. - USA. [read]
Endocrinology. 2020 Dec 29.
Using a reporter mouse to map known and novel sites of GLP-1 receptor expression in peripheral tissues of male mice. Andersen DB, Grunddal KV, Pedersen J, Kuhre RE, Lund ML, Holst JJ, Ørskov C. - Denmark. [read]
Nature. 2020 Dec 16.
Antidepressant actions of ketamine engage cell-specific translation via eIF4E. Aguilar-Valles A, Gregorio D De, Matta-Camacho E, Eslamizade MJ, Khlaifia A, Skaleka A, Lopez-Canul M, Torres-Berrio A, Bermudez S, Rurak GM, Simard S, Salmaso N, Gobbi G, Lacaille JC, Sonenberg N. - Canada, USA. [read]
J Neurosci. 2020 Nov 20.
RFamide-related peptide neurons modulate reproductive function and stress responses. Mamgain A, Sawyer IL, Timajo DAM, Rizwan MZ, Evans MC, Ancel CM, Inglis MA, Anderson GM. - New Zealand. [read]
PLoS Genet. 2020 Nov 2.
A proteomic survey of microtubule-associated proteins in a R402H TUBA1A mutant mouse. Leca I, Phillips AW, Hofer I, Landler L, Ushakova L, Cushion TD, Dürnberger G, Stejskal K, Mechtler K, Keays DA. - Austria, Australia, Germany. [read]
J Clin Invest. 2020 Oct 29.
Defective lysosome reformation during autophagy causes skeletal muscle disease. McGrath MJ, Eramo MJ, Gurung R, Sriratana A, Gehrig SM, Lynch GS, Lourdes SR, Koentgen F, Feeney SJ, Lazarou M, McLean CA, Michell CA. - Australia. [read]
Ozgene PhD graduate
Ozgene staff have an opportunity for further study, either through an external university course or via research at Ozgene. Congratulations to recent Ozgene graduates Maarit Patrick MCom, Liam McMahon LLB, Maya Koentgen BSc (Hons) and Sabrina Koentgen BSc, BA.
Extra special congratulations go to Dr. Maree Hagan, who has been the first to complete an Ozgene PhD in collaboration with Monash University to study Ki-67 protein and its role in immune cell development. Ki-67 is expressed in all dividing cells and assists in chromosome separation during cell division. High Ki-67 expression is most commonly associated with poor cancer prognoses, therefore, improved understanding of Ki-67’s function may lead to the development of new cancer treatments. To address the large gaps in understanding Ki-67 function, Dr. Hagan and the team at Ozgene made a mouse genetically lacking Ki-67. They discovered that Ki-67 was not required for normal life but was specifically required for normal lymphocyte development. Ki-67 expression was linked to successful rearrangement of lymphocyte receptors, indicating Ki-67 has a unique role in immune cell development.
When all processes produce the desired result first time, the timeline can be as short as 18 weeks.
Our fastest conditional KO project took 20 weeks from vector construction to germline transmission.
The current average timeline of our recently completed gene targeting projects is 29 weeks.