Fellow's research: A novel low-cost method to detect chimeric gene fusions in blood cancers


15 May 2020

Fellow's research: A novel low-cost method to detect chimeric gene fusions in blood cancers

Nikhil Patkar, Intermediate Fellow, Tata Memorial Centre, Mumbai

In our latest publication, we describe a novel, low-cost genome sequencing method called NARASIMHA: Novel Assay based on Targeted RNA Sequencing to Identify ChiMeric Gene Fusions in Hematological Malignancies. NARASIMHA aids the diagnosis and treatment of blood cancers driven by gene fusions.

Gene fusions are hybrid genes formed by fusion of two previously separate genes. Chimeric gene fusions (CGF) commonly originate because of chromosomal translocations. About 16% of cancers have a gene fusion as one of their drivers. Gene fusions – considered a hallmark of many cancers – were first detected in the early 1980s. The most widely known and studied gene fusion is BCR-ABL1, which was first discovered as an oncogene in chronic myeloid leukemia (CML). Their accurate characterization is now a critical part of diagnostic and treatment protocols. Traditionally, CGF have been identified in the laboratory using techniques such as karyotyping and fluorescence in-situ hybridization (FISH).

In the last decade, scientists have sequenced organism’s entire mRNA/genes or transcriptomes of thousands of different blood cancers. Identification of novel CGF has opened treatment avenues for these cancers. However, incorporation of CGF testing into routine diagnostics – although urgently required – is possible only through large-scale panels, which are prohibitively expensive and beyond logistic capabilities compared to conventional tests such as FISH and real-time PCR.

Next-generation sequencing (NGS) is an attractive alternative for identification of CGF. However, nearly all the papers that have been published so far suffer from a drawback. They require prior knowledge of both the partner genes involved in the formation of CGF. If the anchor gene in question is promiscuous, there is a strong possibility that the fusion will be missed.

To address this issue, we present NARASIMHA – a cost-effective, laboratory-developed, open-source, targeted RNA sequencing assay for detection of CGF in blood cancers. We named the assay after fiercest mythical avatar of Lord Vishnu – half man, half lion – a chimera. NARASIMHA is based upon a novel element called strand-specific unique molecular motif that marks each DNA strand with a unique molecular fingerprint. This allows for absolute molecule counting and negation of PCR bias. NARASIMHA requires knowledge of only one partner involved in the formation of a CGF and can detect any gene fusion associated with that partner.

In this paper, we describe a modular approach. A 28-gene myeloid or 25-gene lymphoid module is decided on the basis of patient’s pathology results. Between them, they can potentially identify thousands of gene fusions. In our study, we were able to detect 107 CGF (including 5 novel fusions never described in literature) using NARASIMHA, that would be challenging to detect using conventional techniques. These include new disease entities such as BCR-ABL1 like acute lymphocytic leukemia (ALL), B-other ALL, CGF-driven eosinophilia, KMT2A-rearranged leukemia and rare cryptic fusions.  

We describe the clinical, laboratory and treatment outcome features associated with these diseases, and build a case for routine incorporation of targeted RNA sequencing in the workup of haematological malignancies. We estimate that NARASIMHA will make conventional testing such as FISH redundant, accelerating the implementation of precision medicine in haematological malignancies.

Reference:

NARASIMHA: Novel Assay based on Targeted RNA Sequencing to Identify ChiMeric Gene Fusions in Hematological Malignancies. Nikhil Patkar, Prasanna Bhanshe, Sweta Rajpal, Swapnali Joshi, Shruti Chaudhary, Gaurav Chatterjee, Prashant Tembhare, Chetan Dhamne, Maya Prasad, Nirmalya Roy Moulik, Dhanalaxmi Shetty, Anant Gokarn, Avinash Bonda, Lingaraj Nayak, Sachin Punatkar, Bhausaheb Bagal, Manju Sengar, Gaurav Narula, Navin Khattry, Shripad Banavali, P. G. Subramanian & Sumeet Gujral. Blood Cancer J. 10, 50 (2020). https://doi.org/10.1038/s41408-020-0313-6