| 2019 | 2017 | 2016 | 2015 |
Medulloblastoma (MB) is a type of embryonal tumor, meaning it starts in the embryonic cells in the lower back of the brain. It is categorized into four molecular subgroups: Wnt, Shh, Group 3, and Group 4. Out of these four subgroups, Wnt medulloblastoma have the best prognosis as a result of the Wnt activity in the tumor cells. This study focuses on the activation of Wnt pathways in Group 3 and Group 4 MB as a therapeutic treatment for these more aggressive subgroups. Shh MB will not be examined as there is already work showing the antitumorigenic properties of Wnt activation in Shh MB.
Primary patient-derived MB tumor initiating cells were used to create xenografts of tumor tissue for the analytical tests done. First, researchers compared gene enrichment and expression of the sample MB BTIC to bulk MB data, confirming the validity of their sample. It was shown that Group 3 and 4 MB showed higher rates of transcriptional and translational regulation, DNA replication, and RNA processing when compared to the Wnt MB, indicative of a hyperproliferative state. This would explain the aggressiveness of the Group 3 and 4 MB in comparison to the Wnt MB.
Once their samples were validated, the researchers moved into ectopic Wnt activation in the MB BTIC samples. To measure success, they examined three key genes: Bmi1, Sox2, and Axin2. Bmi1 and Sox2 genes both promote proliferation of the tumor cells, and decreased levels of gene expression allude to the effectiveness of the therapeutic Wnt activation. Axin2 is a downstream target gene of the Wnt pathway, useful for indicating Wnt activation. The study found that Wnt activation in Group 3 and Group 4 led to decreases in Bmi1 and Sox2 expression levels and self-renewal levels were greatly reduced.
This study also found that Group 3 and Group 4 MB can sometimes contain rare Wnt-active cells, labeled as TGP+ cells. Similarly, these cells exhibited lower expression levels of Bmi1 and Sox2 genes, lower proliferation, and lower self-renewal when compared with their TGP- counterparts. This further supports the antitumorigenic properties of Wnt activation in Group 3 and Group 4 MBs. This work is crucial to expanding treatment options for Group 3 and 4 MB patients, in which the protective effects of Wnt activation can be applied to these Wnt-inactive subgroups. This study aims to provide a therapeutic pathway that may alter our approach to treating cancer, by reactivating anti-oncogenic cell programs that have been silenced during development.
Links related to the paper:
Wnt: an unexpected tumor suppressor in MB
Deep sequencing of WNT-activated medulloblastomas reveals secondary SHH pathway activation
For animals of different size but of similar anatomy, morphogen gradients play a critical role in scaling of body parts. By having certain segments of a developing organism exposed to certain levels of secreted morphogen, cells will develop differently compared to those exposed to higher or lower levels in the gradient. One such morphogen is SHH (Sonic Hedgehog protein), which affects the neural tube's development. SHH triggers different transcription factors, like OLIG2 (motor neuron cells), NKX2.2 (v3 intermeuron cells) and NKX6.1 (ventral domain cells). It can also be used to repress genes like PAX6 and PAX7 (expressed by GFP) based on how strong the morphogen gradient is. It is not only the type of cell that changes with the morphogen level, but also the size of affected area's tissue (its scale) that is altered. This study specifically tests how SHH gradient and the cells' sensitivity to SHH contributes to the scaling difference between a zebra finch and chick.
Recording size of neural tube tissue collected from developing chick and zebra finch embryos at multiple levels of development, proportion and patterning speed were observed. Expression levels of NKX2.2 and OLIG2 transcription factors were also collected for both species by implanting neural plate tissue with green (NKX) and red (OLIG) dye explants in both and then taking pictures of the explants during certain developmental periods. GFP-expressing chick cells were also implanted into zebra finch neural tubes, and GFP-non-expressing finch cells were implanted into chick neural tubes. After doing so, the NKX6.1, PAX6 and PAX7 levels were recorded from both species to understand morphogen sensitivity. Samples of finch and chick neural plates were collected, stained for NKX2.2 and OLIG2, and exposed to the same level of SHH for multiple different periods of time to further test sensitivity. Finally, more chick and finch samples were given GLI reporter contracts to test GFP expression and GLI activity within both species after testing explants of bothís sensitivity to SAG (SMO protein agent) using RT-PCR.
The resulting data showed that despite the difference in size between zebra finch embryos and chick embryos, the scaling was proportional. The zebra finches, however, finished development into their complete sizes much faster than the chick. The zebra finch also had a lower level gradient of SHH than the chick produced as only OLIG2 and not NKX2.2 (compared to both in the chick) as a result. However, zebra finches cells were much more sensitive to SHH than the chicks and could regulate PAX6, PAX7, and NKX6.1 genes at levels higher than the chicks, which explains their fast patterning time even with lesser SHH levels. Further proving sensitivity, chicks had lesser NKX2.2 and OLIG2 expression when exposed to equal amounts of SHH, as well as being less sensitive to SAG and showing SMO's downstream mechanisms were what was causing the sensitivity discrepancies. GLI3 in the neural tube was shown to be the transcription factor with different levels of expression between the species and the ultimate cause of patterning differences.
Links related to the paper:
Neural Tube Development and SHH
Morphogen Gradients and Scaling
One of the leading types of cancer that causes death in humans every year is related to different forms of non-small lung cancer, and specifically lung adenocarcinoma. Lung adenocarcinoma has been identified as one of the many forms of cancer that contains a large number of copy unknown alterations and mutations within the tumorsí genomes, which leads to the high fatality rate in many patients. A thorough analysis of these mutations proved to impact key pathways within organisms, one of which being the receptor tyrosine kinase (RTK) pathway. Due to the many unknown mutations, researchers use model organisms, specifically mice in this experiment, to determine in vivo the tumorigenicity of the specific mice genes that cause lung adenocarcinoma, so that the human ortholog genes for lung adenocarcinoma can be more widely understood. This information and greater knowledge are then used to target genes that can act as tumor suppressors when influenced by mutated KRas and the RTK pathway.
Locating specific genes that contain tumor suppressor activity towards lung adenocarcinoma were first targeted by using an shRNA-mediated high-throughput approach. This shRNA approach allowed for a screening test to be implemented into mutant KRasG12D mice. KRasG12D mice develop benign tumors when independent of other mutations, but when combined with other genome mutations, it cooperates with other genes to delete specific tumor suppressor genes in mice leading to diseases like lung adenocarcinoma. When KRas is mutated, it transforms from a proto-oncogene to an oncogene. The shRNA screen in the KRasG12D mutants identified ephrin receptor A2 (EphA2) as being a main signal transduction pathway gene that also inhibits tumor growth and the spread of lung adenocarcinoma. EphA2 is a member of the Ephrin receptor family that is directly involved in the RTK pathway. Further experiments that silenced and knocked down this EphA2 gene through the creation of shEphA2 or shControl cell lines in mutant mice produced large levels of cell proliferation, and little to no suppression or regulation. This lack of suppression leads to the RTK pathway, specifically KRas, to promote lung adenocarcinoma. These different shRNA screens also showed that this target gene, EphA2, can be used in therapeutic treatments in patients with lung adenocarcinoma due to its tumor-suppressing capability.
The results of this study offer wider knowledge about different target genes that can be used to help treat patients with lung adenocarcinoma, and even potentially other forms of non-small cell lung cancer. By identifying the effect that EphA2 had on tumor suppression in KRasG12D mutant mice, researchers could also identify the main signal transduction pathways, like the receptor tyrosine kinase (RTK) pathway, that lead to increased cell proliferation and tumorigenesis. This paper briefly discusses the impact that EphA2 loss has on the hedgehog signaling pathway as well, but further research should be done on subsequent signal transduction pathways to gain a greater understanding of tumor suppressor target genes, and the unknown mutations and alterations that create oncogenic genes that promote lung adenocarcinoma.
Links related to the paper:
EphA2 and the Ephrin Receptor Family
What is shRNA or High-throughput Screening?
Common RTK Mutations in Non-small Cell Lung Cancer
The purpose of this study is to create a method for screening potential driver mutations of acute myeloid leukemia using a cDNA library and transplantation experiments in mice. Acute myeloid leukemia, or AML, accounts for half of the leukemia cases in teenagers and most cases in adults. This form of leukemia takes place when myoblasts reproduce without developing into normal blood cells. AML starts in the bone marrow and comes on very suddenly compared to other forms of leukemia. This study worked to determine genes and potential combinations of genes causing leukemia to aid in leukemia therapy.
In this study, murine bone marrow cells were transduced with oncogenes and homeobox genes, found in the retroviral cDNA library, and transplanted into mice. Initially, different combinations of genes including MYC, KRAS, and PIM2 were tested. This was not conclusive because MYC is known to generate leukemia independently. For further investigation, MYC was removed from the cDNA library and the MYC-deleted cDNA library was transplanted into mice. This induced acute myeloid leukemia in all mice within 6 to 15 weeks.
The mice were divided into four different groups. The first group had MEIS1 and HoxA6 as their main cDNA inserts, the second group had MEIS1 and HoxB6, the third group had MEIS1 and HoxB7, and the fourth group had MEIS1 and HoxD8. The results of these trials showed HoxB7 and HoxD8 as potential genes acting with MEIS1 to induce AML. After testing both of these combinations, it was clarified that the co-transduction of MEIS1 with HoxB7 or HoxD8 were able to induce leukemia, but none of these genes were able to induce leukemia alone.
This system or model generated in this study is more efficient than previous models. However, future investigation is necessary to resolve technical issues in the process.
Links related to the paper:
Defining Roles for HOX and MEIS1 Genes in Induction of Acute Myeloid Leukemia
Make and Screen a cDNA Library
Animal Models Used in AML Pathophysiology
The Sonic Hedgehog gene is vital in the role of embryonic development in many animals and for tumor suppression. If Shh is downregulated then it results in birth defects such as Holoprosencephaly. Holoprosencephaly can be present in varying degrees: alobar, semilobar, or lobar. Alobar is the most severe, resulting in the lack of division in the brain (no lobes) and is associated with severe facial deformities. Semilobar is when the brain is partially divided and is the intermediate form of the birth defect. Lobar is the least severe and is the least likely to be fatal. The brain, in some cases can appear to be normal. In other cases, there is substantial evidence that the brain has separate hemispheres. The misregulation or uncontrolled regulation of Shh can cause cancer, such as Golin Syndrome. Golin syndrome is also known as nevoid basal cell carcinoma, meaning that there is a higher risk of the person developing skin cancer throughout their life, but with proper care can be manageable. In unstimulated cells the activity of Shh is inhibited by the Patched1 gene. In this study the mechanism by which Shh inhibits or antagonizes Patched1 was investigated in mice.
Palymitoylated Shh22 was tested to verify its potency in the induction of transcription Gli. Palmitoylated Shh22 was added to the N terminus of a bacterial HaloTag to track where it was being expressed in signaling. To verify that Shh22 was not activating Shh and therefore being upregulated as a result of the presence of Shh22, the 5E1 monoclonal antibody was used to bind to Shh and inhibit it, but does not bind to Shh22. Shh22-NanoLuc was also used to verify that the HaloTag was not causing any novel activity in the signaling pathway. Assays were performed on the initial steps of Hh signaling to verify that Patched1 was inhibited and that Smo was accumulating in the cilia, where the initial steps of Hh take place in vertebrates.
The results showed that together the palmitoylated N terminus of Shh is sufficient for Ptch1 inhibition and for triggering Hh signaling and that the rest of Shh is not absolutely required. The inhibition of Patched1 is palmitate-dependent. The palmitate-dependent interaction between Shh and Patched1 results in holoprosencephaly. Patched1 point mutations, however, abolish the interaction and cause Gorlin cancer syndrome.
Links related to the paper:
Sonic hedgehog signaling by the Patched-Smoothened receptor complex
Palmitoylation of Sonic Hedgehog
Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and is the second most common cause of cancer-related death worldwide. Targeting the Wnt/β-catenin pathway as a therapy for HCC is promising because the activation of this signaling pathway leads to cell survival and proliferation and is a frequent event in this type of cancer. Upon activation of Wnt signaling, β-catenin accumulates and translocates into the nucleus and forms a transcriptionally active complex, two coactivators of the β-catenin transcription factor complex, BCL9 and BCL9L, result in the activation of downstream target genes.
HCC cell lines and normal cell lines were cultured and analyzed. BLC9 and/or BCL9L were knocked out with siRNAs. Quantitative real-time PCR measured mRNA expression levels, while western blot analyzed proteins by using antibodies against BCL9 and β-catenin. The viability and apoptosis of both the cell lines were also measured. Wnt/β-catenin signaling activity was assayed to group the cells into Wnt-inactive and Wnt-active cells.
This study found that the knockdown of BC9L, but not BCL9, reduced Wnt-signaling activity and reduced cell viability and increased apoptosis of Wnt-inactive cells, not Wnt-active cells. Expression of BCL9L was observed in larger tumors and correlated with poor overall survival. This study suggests that BCL9 and BCL9L are not limited to their known role as cofactors in the β-catenin transcription complex since they appear to function independent of the Wnt-signaling pathway and may have more unknown functions. Directly targeting these component proteins of the Wnt-signailing pathway seem to be potential target therapies to inhibit HCCs.
Links related to the paper:
Hepatocellular Carcinoma: Overview and Risk Factors
The Role of Wnt-Signaling in Cancer
Hepatocellular Carcinoma Cell Lines as in vitro Models
C. elegans has an Insulin like signaling pathway that plays a part in regulating overall longevity in C.elegans. It has also been observed that the transcription factor DAF-16/FOXO initiates dauer development. In order to find out more information, they looked into the mirroring homolog SKN-1 that can be inhibited by the Insulin signaling pathway
Based on the results, it has been observed that the SKN-1 increases the expression of collagen and other ECM genes (remodeling) which contributes to increased longevity when Insulin like signaling pathway is blocked. Collagens are needed in order to slow the aging process by interventions that don't involve dauer characteristics. It has also been revealed that the Insulin like signaling pathway is also in charge of producing protective mechanisms in regards to longevity inn C.elegans.
Links related to the paper:
Cytoprotective perspective on longevity regulation. Trends in cell biology.
Dauer larvae and long-lived daf-2 mutants implicates detoxification system in longevity assurance.
Direct inhibition of the longevity-promoting factor SKN-1 by insulin-like signaling in C elegans
While genes that support embryonic development are essential for proper functions, many are not needed in adult organisms. When such genes arise in cells there can be negative effects on the organism. This is observed between the LIN28 gene and colorectal cancer. The LIN28 gene, unlike other genes observed in intestinal colorectal cancer, can be targeted because it is not a needed gene within normal adult intestines. This means research to prevent or slow down colorectal cancer can be directed toward LIN28 because if it is inhibited there are no negative effects on adult intestines. LIN28A and LIN28B are paralogs found in the early development of mammals. They are not meant to be expressed within adult mammals particularly in high levels. As their levels decrease the levels of let-7 microRNA, regulated by LIN28 increases. However, when either LIN28 paralog is over-expressed it can interact with the WNT signaling pathway to promote cancerous cells and block let-7 miRNA. LIN28 interacts with the Wnt signaling pathway through a beta- catenin mutation. They both encourage the invasion of intestinal adenocarcinoma and LIN28 assisted in undifferentiated cells.
The roles of LIN28A and LIN28B are observed in intestinal epithelium in mice. Different experiments are conducted to compare the two LIN28 paralogs to each other, how both relate to other mutations, their significance in different mice, and its overall effects. Several experiments involved testing LIN28A and LIN28B separately to determine if they are expressed in the same manner and to the same degree. These were conducted with transgenic mice. One set of mice contained the over-expressed intestine LIN28A gene from mice and referred to as iLin28a. Another set of mice had the over-expressed LIN28B gene from human intestines and were referred to as iLIN28B. Other experiments used the transgenic mice to observe the effects of each LIN28 paralog and differentiate their effects while also studying their individual reaction. Another set of mice used are APCmin/+ where the tumor suppressor gene, APC no longer functions causing the formation of several intestinal tumors. These mice are used and tested for LIN28 and compared with iLin28a and iLIN28B to observe the different tumors.
Each experiment was meant to support the hypothesis that LIN28 encourages the continuation of undifferentiated cells in intestinal cancer. This study contributes to the search for a cure or treatment to colorectal cancer. The devastation this cancer causes is highly significant which is why there is so much research relating to different genes and pathways. Each experiment, hypothesis, and research paper lead to a solution.
Links related to the paper:
Much of what is known regarding the initiation and regulation of the Hox gene lin-39 has been studied and researched on primary vertebrates and in Drosophila. The information and data that we have currently gathered for the Hox gene lin-39 regulation in organisms such as C. elegans are very limited. Especially when we come across mutations that are related to lin-39 expression, which is required for correct fate specialization in the mid-body region (including the VPCs), it becomes critical to understand why these mutations happen and it is similar to Drosophila and vertebrates.
By using the yeast one-hybrid (Y1H) methods, we are able to assess DNA fragment sequences on whether the necessary protein factors drive and regulate lin-39 expression in the development of the vulval precursor cells. This technique was performed, both, traditionally and robotically.
The results suggest that the necessary protein factors, that were tested in C. elegans, positively regulated levels of lin-39 and lin-39::GFP. These protein factors included NHR-43, LIN-26, ELT-6, ZTF-17, BED-3, and TBX-9. In addition, the results also continue to suggest that there could be other potential factors that affect lin-39 expression in the VPCs that hasnít been researched yet.
Links related to the paper:
Yeast one-hybrid system (Y1H) simple, brief, and complete
C. elegans hox gene lin-39 controls cell cycle progression during vulval development
Comparsion of the nematode Hox gene lin-39 in C. elegans and P. pacificus
Chordoma is a rare tumor found at the base of the skull or spinal cord that forms from remnants of embryonic notochord. Many mutations are suspected to be the cause of overproliferation of notochord cells but, prior to this experiment, no in vivo evidence was available. Brachyury is a notochord regulator protein that is expressed in chordoma. In this experiment, brachyury is tested as a possible causative agent of chordoma formation.
An injection-based assay was used to run the experiment, following a Gal4/UAS system. The transgene col2a1aR2:KalTA4 was used for injected transgene expression in the notochord. Because Brachyury was suspected to be the source of initiating notochord hyperplasia, both human and zebrafish brachyury genes were tested and both came back with normal notochord formation. Notochord hyperplasia was only triggered when chordoma-related RTK genes EGFR and Kdr/VEGFR2 were injected. RTK/Ras-transformed notochords of zebrafish were sequenced and the results showed a misexpression of human chordoma-related genes and a deregulation of unfolded protein response (UPR).
The results suggest that abnormal RTK signaling pathway is responsible for initiating notochord hyperplasia that leads to chordoma formation. This could be due to the overexpression of RTK genes that signal chordoma formation and early notochord formation that causes stress to the Endoplasmic Reticulum(ER) and the unfolded protein response(UPR). Further findings from the experiment suggest that brachyury is not responsible for hyperplasia of the notochord. In contrast, the RTKs, EGFR and Kdr/VEGFR2 were found to be responsible for transforming the notochord cells and initiating chordoma formation.
Links related to the paper: