Gefitinib

Lung adenocarcinoma in a patient with Li–Fraumeni syndrome bearing a novel germ-line mutation, TP53R333Vfs∗12

Shodai Takahashi1,†, Kazuhiro Shimazu1,†, Koya Kodama2, Koji Fukuda1, Taichi Yoshida1, Daiki Taguchi1, Tsutomu Takahashi2, Hiroshi Nanjyo3 and Hiroyuki Shibata1,*
1 Department of Clinical Oncology, Akita University, Akita, Japan,
2 Department of Pediatrics, Akita University, Akita, Japan and
3 Department of Pathology, Akita University Hospital, Akita, Japan

Abstract

Germline mutations of TP53 are responsible for Li–Fraumeni syndrome in its 60–80%. We found a novel germline mutation, TP53: c.997del:p.R333Vfs∗12 (NM_000546.6, GRCh, 17:7670713..7670713). The proband is a 40-year-old female, who was suffered from osteosarcoma in her right forearm at her age of 11. She was also suffered from lung adenocarcinoma in her right upper lobe and bone metastasis in her right scapula at her age of 37. She was treated with gefitinib, an epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) because of EGFR mutation (L747-S752 del). Her bone metastasis became resistant after 1-year treatment. Bone metastasis had an additional EGFR mutation (T790M). The secondary treatment with osimertinib, an another EGFR-TKI, can successfully control the tumors for over 2 years. This TP53 mutation (R333Vfs∗12) was first found in lung adenocarcinomas. The therapeutic effect of osimertinib for this triple mutant lung adenocarcinoma is better than the previous report.

Key words: TP53, epidermal growth factor receptor-tyrosine kinase inhibitor, lung adenocarcinoma, Li–Fraumeni syndrome

Introduction

Li–Fraumeni syndrome (LFS) is an autosomal dominant predisposi- tion to various malignancies including sarcoma, glioma and breast cancer. Clinically, LFS is diagnosed by three criteria: (i) sarcoma before age 45, (ii) malignancy in first-degree relative before age 45 and (iii) malignancy in another first- or second-degree relative before age 45 or sarcoma regardless of age. Germline TP53 gene mutations are most frequently responsible for LFS. Loss of function of TP53 protein results in alterations in DNA repair, apoptosis cell cycle control and consequent genome instability.
The newest version (R20) of TP53 database in International Agency for Research on Cancer (IARC) lists 29 900 types of somatic mutations and 1530 germline mutations in LFS families (https://www.iarc.fr/news-events/new-release-of-the-iarc- tp53-database-2019/). The prevalence of lung cancer is rather rarer and accounts for approximately 25% of LFS (1). The Cancer Genome Atlas indicates that TP53 mutations were identified in 46% of sporadic lung adenocarcinomas (LAC) (2).
In patients with lung cancer, the TP53 status is not currently used in therapeutic decision-making (2). Immunohistochemistry indicated that TP53-positive staining conferred inferior survival (2).
Co-incidence of TP53 mutation and epidermal growth factor receptor (EGFR) mutation was reported to be 62% (3). There was no significant difference in progression-free survival (PFS) compared with the subgroup of TP53 mutation alone treated with EGFR-tyrosine kinase inhibitors (EGFR-TKIs). Oppositely, patients with double mutations of EGFR T790M and TP53 had worse survival compared with patients with T790M mutation alone (46.4 vs. 82.9 months) (3). It is still unknown if all TP53 mutants confer clinical outcomes.
In this report, the therapeutic outcome of LAC bearing a new TP53 mutation, TP53: c.997del:p.R333Vfs∗12 (NM_000546.6, GRCh, 17:7670713..7670713), in a patient with LFS is described.

Case report

A 41-year-old index female patient (II: 2) was considered to have LFS after her son (III: 1) was diagnosed with rhabdomyosarcoma and died at 6 years (Fig. 1). She was diagnosed with osteosarcoma at age 11 (1990) second in her pedigree after her father (I: 1), who was diagnosed with osteosarcoma in childhood and died from gastric cancer at the age of 42 (Fig. 1). The osteosarcoma in her right humerus was resected and replaced with fibula. After perioperative chemotherapy with methotrexate and vincristine (total nine cycles), her osteosarcoma was cured. After her son’s onset, a novel germline mutation of R333Vfs∗12 was discovered in her germ line as well as her son’s (4). Genomic DNA of peripheral blood leukocyte from the index patient, her son as well as her husband was examined by trio whole-exome analysis (4). This mutation was considered to be a pathogenic mutation from its structural change and its genetic inheritance. R333Vfs∗12 was never reported in the TP53 database (R20) in IARC at that time. At 37 years (2016), she felt right shoulder pain. After whole body examination, right lung tumor, bone tumor in her right scapula, thyroid tumor and uterine tumor were apparent (Fig. 2A and B). Endoscopic cytological examination revealed that her lung tumor was adenocarcinoma bearing an EGFR mutation, L747–S752 del. Thyroid and uterine tumors were benign, and bone tumor was a metastasis from LAC. She visited our department in October 2016 and was treated with gefitinib. Gefitinib controlled the LAC for 313 days. The best response was partial response (PR), judged by shrinkage of the tumor of the largest diameter from 40 to 20 mm (Fig. 2A and C). The adverse events were mild except grade 3 paronychia. However, right scapula destruction advanced after treatment for 1 year (Fig. 2D), and the bone metastasis became resistant to gefitinib. A bone biopsy was conducted, which found TTF1-, Napsin A- and TP53-positive adenocarcinoma (Fig. 3), where acquired resistant mutation T790M was detected. Due to these findings, we changed gefitinib to osimertinib, a third-generation EGFR-TKI. Further, radiotherapy (30 Gy/10 fractions) was added to this lesion, and denosumab, an anti-receptor activator of nuclear factor-κB ligand antibody, was started to prevent skeletal-related events such as bone fracture. Since then, primary and bone metastases have been under control with osimertinib without any severe adverse events for more than 781 days. The best response was stable disease (Fig. 2E and F). There is no apparent carcinogenesis in the other sites.

Discussion

Patients with LFS face the risk of malignancies throughout their lives. Previously, the life expectancy of LFS patients experiencing early-onset malignancies was estimated to be <40 years (5). Recent advances in cancer chemotherapy might contribute to patient sur- vival. Elongation of survival time demands attention to carcinogene- sis in organs beyond the classic LFS-related malignancies. The TP53 germline mutation, R333Vfs∗12, in a LFS pedigree has never been reported in IARC TP53 database. R333C was detected in Australia, in two females with acute leukemia and Ewing sarcoma, respectively. R333G was detected in a female of unknown nationality, with hepatocellular carcinoma. We analyzed transcriptional activities of the missense mutations at codon 333, located in the tetramerization domain of TP53 (6), which resulted in loss of TP53. The germline mutation most similar to ours in IARC TP53 database is a one base pair (bp) deletion at codon 333 (c. 997_998) detected in a Chinese family. The female proband had breast cancer. This frameshift muta- tion in tetramer domain results in loss of tetramerization of TP53. In the cases of somatic mutations at codon 333, five cases were reported, including three cases of 1 bp deletion in ovarian cancers in USA, Poland and an unknown nation. One case of 2 bp deletion and one case of 14 bp insertion at codon 333 were detected in breast cancers of females in the USA and Sweden, respectively. In the COSMIC database (https://cancer.sanger.ac.uk/cosmic), R333Vfs∗12 TP53 mutation were reported from three cases of breast cancer, three cases of prostate cancer, two cases of melanoma, one case of colon cancer and one case of brain tumor. TP53R333Vfs∗12 mutation could be a loss-of-function mutation, given that two-thirds of its tetramerization domain (codon 325–355) and its C-terminus are lost. Reports from the literature of missense mutations in this domain support this (7). Loss of the tetramerization domain does not lead to a classical dominant-negative gain of function. Haploinsufficiency of TP53 could be a cause of carcinogenesis in LAC, as has been shown in a mouse model; however, we did not analyze this (8). In the Akita pedigree bearing this mutation in their germ lines, two cases of osteosarcoma, one case of rhabdomyosarcoma, one case of gastric cancer and one case of LAC were found (4). Detections of R333Vfs∗12 TP53 mutation from these tumors are the first reported in the literature. The EGFR mutation, L747–S752 del, existed in the LAC, and the second mutation, T790M, occurred after 313 days of treatment with gefitinib. Osimertinib, another EGFR-TKI, can successfully control the LAC over 781 days. Lung cancer accounts for around 25% of LFS; however in the sporadic lung cancer, TP53 mutations are detected in 46–81% (9). EGFR mutation rate in non- small cell lung cancer (NSCLC) was highest (47%) in countries in Asia (9). Median PFS (mPFS) of gefitinib for EGFR mutant-NSCLC is 10.4 months (10). Our patient responded to gefitinib similarly. The second site EGFR T790M mutation was detected in 48–62% of LAC pre-treated with first-generation EGFR-TKI (11). mPFS of osimertinib for pre-treated EGFR T790M-positive advanced NSCLC was 9.9 months (11). Our patient with LAC was under control for over 27 months. It is the single report of EGFR-mutated LAC in LFS, where all of six cases have TP53 germline mutations in the DNA-binding domain. They were treated with the first- or second-generation EGFR-TKI, and the best responses were one complete response, three PRs, one progressive disease and one unspecified (12). LAC in patients with LFS might respond well to EGFR-TKI. EGFR T790M mutation was observed in only one case, but this patient was not treated with osimertinib. On the other hand, Aggar- wal reported that median overall survival (OS) in both TP53 and EGFR T790M mutant-NSCLC (46.4 months) was worse than that in TP53 wild but EGFR T790M mutant-NSCLC (82.9 months), treated with EGFR TKI including osimertinib (3). The mPFS in both TP53 and EGFR T790M mutant-NSCLC was 15.9 months. However, only 7% of TP53 mutations occurred in exon 10, where the tetramerization domain locates. Perhaps, all of them are not LFS. Furthermore, Fischer reported that the median OS in subjects with missense mutations in the tetramerization domain, which potentially forms multimeric TP53 (51 months), was better than those with mutations in the tetramerization domain, which potentially forms monomeric TP53 (33 months) (7). The difference between these two groups was very large. This study indicated that not all TP53 mutations are equivalent. 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