Emory Klann
In their recently published research paper, Kulsom Nuuri and Dr. Yoana Arroyo-Berdugo from the University of Roehampton presented their investigation of the origin of chemotherapy resistance in pediatric patients suffering from acute lymphoblastic leukemia (ALL). In ALL, large numbers of lymphoblasts develop from stem cells in the bone marrow and quickly crowd out other blood cells that are important for fighting infection, transporting oxygen, and clotting. Eventually, these cancerous cells spread to the blood and other organs. Without treatment, ALL can be fatal within just a few months.
Image by: James Grellier - derivative of original work by VashiDonsk at en.wikipedia / CC BY-SA (https://creativecommons.org/licenses/by-sa/3.0)
The onset of ALL is relatively rapid, but there is a high chance of survival, especially in children. However, leukemias in 60-70% of infants are associated with a rearrangement of the Mixed Lineage Leukemia (MLL) gene, and the survival rates for these patients are poor. MLL is an important gene regulator, and sometimes it can fuse with a partner protein. The fusion of MLL and AF4 is the most common MLL translocation in ALL, present in the SEM-1 human cell line and is found in 80% of leukemias. The fusion of MLL and ENL is present in the human cell line HB-1110. Both MLL-AF4 and MLL-ENL are capable of inducing ALL. This occurrence is linked to resistance against one of the most common treatments for pediatric ALL, making it a crucial research focus.
To further evaluate chemotherapy resistance in MLL-ALL, researchers examine two genes of the DNA mismatch repair (MMR) pathway, MSH2 and MSH3, within cell lines that are resistant to chemotherapy. These genes are important for the formation of MutS proteins. These bind and recognize mismatch errors, initiating the MMR pathway.
Tumor cells that are deficient in MSH2 are more vulnerable to the generation of reactive oxygen species (ROS) and other changes that may eventually lead to apoptosis, or programmed cell death. Silenced MSH2 genes in another form of leukemia, acute myeloid leukemia (AML) showed increased sensitivity to treatment drugs. This information prompted researchers to analyze the roles of MMR genes of ALL cells in chemotherapy resistance.
The researchers focus on two groups of cell lines: the non-MLL (control) cell lines Kasumi-1 and K562, and the experimental cell lines HB-1119 and SEM-1 cell lines, which both contain MLL genes fused with partner genes. All four of these were cultured in vitro, and cDNA was generated from the isolated RNA via Retro-transcription-Polymerase Chain Reaction (RT-PCR). Then endpoint and semi-quantitative PCR were used to analyze gene expression.
Endpoint PCR showed that MSH2 and MSH3 were expressed in all four cell lines, but semi-quantitative PCR indicated that expression of MSH2 was higher than MSH3 in Kasumi-1 and HB-1119, and MSH3 expression was higher in SEM-1 and K562. The lower expression of MSH2 in HB-1119 and Kasumi-1 demonstrates possible microsatellite instability in MLL-ALL, but additional research is necessary to determine the threshold of MSH2 necessary to induce this.
Although MSH2 expression was higher in Kasumi-1 relative to MSH3 expression, both genes were expressed. Kasumi-1 is sensitive to chemotherapy treatments, thus this data reveals the possibility that deficiency of MMR genes is not necessary for treatment sensitivity.
Both MSH2 and MSH3 were downregulated in HB-1119 in comparison with SEM-1. Therefore, it may be more likely to be MMR-deficient. However, it is important that further research identifies the proteins that can compensate for the downregulation of MSH2 and MSH3 in these cell lines to find if the MMR pathway is completely deficient. The downregulation of MSH2 in HB-1119 indicates that this cell line might be more vulnerable to chemotherapy treatment toxicity. On the other hand, the downregulation of MSH3 in the same cell line reveals that HB-1119 could show resistance.
The outcome of this research alone demonstrates that the expression of a single gene is not ample enough to evaluate treatment resistance, but this is a positive start. With further extensive research on MMR’s range of capabilities, as well as knockdown and overexpression studies to find each gene’s individual role, it is possible to recognize methods as means of uncovering the vast range of duties that specific genes have in chemotherapy resistance. Nuuri and Dr. Arroyo-Berdugo’s research has the potential to launch the advancement of leukemia research so that one day, more and more children will survive ALL.