Molecular tumor board: molecularly adjusted therapy upon identification and functional validation of a novel ALK resistance mutation in a case of lung adenocarcinoma.

We report a case of a long-term surviving patient with EML4/ALK translocated non-small cell adenocarcinoma of the lung in UICC8 stage IVA. During recurrence under continuous crizotinib therapy, a hitherto insufficiently characterized missense mutation in the ALK gene (Arg1181His) was identified through targeted sequencing. The aforementioned EML4/ALK translocation could still be detected in this situation. Employing a 3D reconstruction of the ALK tertiary structure, considering its interaction with various ALK inhibitors at the molecular binding site, our analysis indicated the presence of a mutation associated with crizotinib resistance. To validate the biological relevance of this previously unknown mutation, we carried out an in vitro validation approach in cell culture in addition to the molecular diagnostics accompanied by the molecular tumor board. The tumor scenario was mimicked through retroviral transfection. Our comparative in vitro treatment regimen paired with the clinical trajectory of the patient, corroborated our initial clinical and biochemical suspicions. Our approach demonstrates preclinical, in silico, and clinical evidence of a novel crizotinib resistance mutation in ALK as well as sensitivity toward brigatinib and potentially lorlatinib. In future cases, this procedure represents an important contribution to functional diagnostics in the context of molecular tumor boards.

Late Onset of Primary Hemophagocytic Lymphohistiocytosis (HLH) with a Novel Constellation of Compound Heterozygosity Involving Two Missense Variants in the PRF1 Gene.

Hemophagocytic lymphohistiocytosis (HLH) is a rare but in most cases life-threatening immune-mediated disease of the hematopoietic system frequently associated with hematologic neoplasms. Here, we report on a case in which we detected a novel constellation of two missense variants affecting the PRF1 gene, leading to de novo primary HLH. Diagnostics included a comprehensive clinical work-up and standard methods of hematopathology as well as extended molecular genomics based on polymerase chain reaction (PCR) reactions and the calculation of three-dimensional molecule reconstructions of PRF1. Subsequently, a comprehensive review of the literature was performed, which showed that this compound heterozygosity has not been previously described. The patient was a 20-year-old female. Molecular diagnostics revealed two heterozygous missense variants in the PRF1 gene (A91V and R104C) on exon 2. Apart from the finding of two inconclusive genetic variants, all clinical criteria defined by the HLH study group of Histiocyte Society were met at initial presentation. The final diagnosis was made in cooperation with the Consortium of German HLH-reference centers. Here, chemotherapy did not lead to sufficient sustained disease control. Therefore, the decision for allogenic hematopoietic stem cell transplantation (alloHSCT) was made. Hitherto, the duration of response was 6 months. Due to severe and unmanageable hepatic graft-versus-host disease (GvHD), the patient died. We report on a novel constellation of a compound heterozygosity containing two missense variants on exon 2 of the PRF1 gene. To the authors' best knowledge, this is the first presentation of a primary HLH case harboring this genomic constellation with late-onset clinical manifestation.