Primary Mutations Research Articles

Practical Implications of KRAS Mutation Status and Sidedness of Primary Tumour in Patients with Colorectal Cancer and Synchronous Liver Metastases: A Subset Analysis of the CoSMIC Study.

Simple SummaryPatients presenting with liver metastases at the time of diagnosis of their colorectal cancer (termed ‘synchronous disease’) have a worse outcome than those whose disease is limited to the primary bowel tumour. There is evidence to show that patients with tumours on the right side of the colon have worse survival. Furthermore, mutations in the KRAS gene have also been shown to adversely affect outcome. This study explores the association between the side of the primary colorectal tumour in patients with synchronous disease and mutations in the KRAS gene on survival. We analyse a specific cohort of patients from the previously published CoSMIC study who presented with colorectal cancer and synchronous liver metastases and who had their colorectal tumour analysed for KRAS mutations. We find that for this cohort, neither mutations in the KRAS gene nor the side of the colon tumour influenced survival.Patients with colorectal cancer presenting with synchronous liver metastases have less favourable outcomes than those with primary-only disease. There is evidence of different genetic mutational signatures according to the sidedness of the primary tumour. KRAS mutations are key driver mutations in colorectal cancer progression. This post hoc analysis of the previously reported CoSMIC inception cohort explores the association between primary tumour sidedness and KRAS mutational status on the outcome of patients with colorectal cancer and synchronous liver metastases. Patients diagnosed with synchronous disease were recruited between April 2014 and March 2017 and, after exclusions, 83 patients undergoing colorectal primary KRAS mutation testing constituted the final study population. Data were collected prospectively on demographic profiles, treatment, and outcomes. Twenty-one patients (25%) had right-sided tumours and 62 (75%) had left-sided tumours, with 46 (55%) and 37 (45%) exhibiting wildtype and mutated KRAS, respectively. There was no difference in distribution of liver metastases by KRAS status (unilobar vs. bi-lobar; p = 0.58; Fisher’s Exact test) and no difference in 5-year survival according to KRAS mutation status (Log-rank test, p = 0.82) or tumour sidedness (p = 0.16). In summary, in this cohort of patients with colorectal cancer and synchronous liver metastases, neither KRAS mutation status nor tumour sidedness influenced survival.

64 (PB054) - BDTX-1535, a fourth generation EGFR inhibitor, targeting intrinsic and acquired resistance mutations in NSCLC

Background: The classical mutations of the epidermal growth factor receptor (EGFR) in NSCLC are the exon 19 del and exon 21 (L858R) mutations. The 3rd generation EGFR tyrosine kinase inhibitor (TKI), osimertinib, has become the first-line treatment of choice for NSCLC patients with these mutations. Resistance to 3rd generation TKIs can be driven by gain of secondary EGFR alterations, e.g., substitution at cysteine-797 to serine (C797S). Real world evidence shows that in addition to C797S, other EGFR alterations are also acquired upon osimertinib treatment, including kinase domain mutations (e.g., S768I), extracellular domain alterations (e.g., EGFRvIII, A289X), and EGFR amplification. In addition to the classical mutations, NSCLC tumors express a wide spectrum of primary kinase domain mutations including G719X in exon 18, S768I in exon 20, and L861Q in exon 21, which confer intrinsic resistance to approved TKIs. NCCN guidelines recommend the use of afatinib or osimertinib, however, there remains a need for an EGFR TKI that targets these mutations with high potency, good tolerability, and CNS penetration. Our MAP platform allowed us to select MasterKey inhibitor BDTX-1535, a 4th-generation CNS penetrant EGFR TKI, that potently and selectively targets multiple intrinsic and acquired resistance EGFR alterations. Materials and Methods: BDTX-1535 preclinical exposure was evaluated across species, and PK and Kpuu values were calculated in brain and plasma. Antitumor activity was assessed across a broad range of mouse PDX and allograft models. Results: BDTX-1535 is a potent and selective, CNS penetrant, wild type sparing, irreversible EGFR TKI targeting Exon 18–21 alterations which are associated with intrinsic or acquired resistance to 3rd generation EGFR inhibitors. These include a broad range of kinase domain EGFR mutations (e.g., C797S, L718Q, G724S, S768I), extracellular domain alterations (e.g., EGFRvIII, A289X), and EGFR amplification. Mouse xenograft and allograft studies showed that BDTX-1535 consistently achieves regression of tumors carrying these mutations at well tolerated doses without significant EGFR wild type associated toxicities. BDTX-1535 is orally bioavailable with a CNS Kpuu of 0.55 and 0.48 in rat and dog, respectively, and active in an intracranial PDX model. Conclusions: BDTX-1535 is a 4th generation CNS penetrant EGFR TKI discovered using the MAP platform which allows MasterKey targeting of EGFR alterations associated with intrinsic or acquired resistance to 3rd generation EGFR TKIs, while sparing wild type EGFR. This broad spectrum coverage of resistance mutations in conjunction with CNS penetration properties allows BDTX-1535 to address a key unmet medical need in EGFR mutant lung cancer. BDTX-1535 is currently under phase I clinical investigation (NCT05256290). Conflict of interest: Ownership: Black Diamond Therapeutics

Primary resistance against integrase strand transfer inhibitors in integrase strand transfer inhibitor-naive patients failing first- and second-line ART in Tanzania.

Sub-Saharan African countries are introducing integrase strand transfer inhibitors (INSTIs) in their ART programmes as the preferred first-line regimen, and dolutegravir is the INSTI of choice due to its potency, tolerability and high genetic barrier to resistance. Dolutegravir was introduced into the first-line ART regimen in Tanzania in 2019. However, there is a paucity of data on the occurrence of mutations in HIV lineages circulating in Tanzania. This study aimed to determine the prevalence of INSTI primary resistance mutations in Tanzanian patients exposed to ART but not INSTIs. Plasma samples from 50 INSTI-naive patients failing first- or second-line ART [median (IQR) age: 40 (21.93-46.41) years; 68% women] were subjected to Sanger sequencing of the HIV integrase gene. Participants had been on ART for a median (IQR) duration of 7.32 (4.73-9.29) years, with 80% and 20% failing first- and second-line ART, respectively. No major INSTI mutations were found, but 2 (4%) participants had the accessory mutation T97A. Using the REGA HIV-1 subtyping tool, HIV subtype A1 (53.1%) was found to be dominant, followed by subtypes C (30.6%) and D (16.3%). This study found no current evidence for transmitted resistance against INSTIs among unexposed patients failing ART and supports the scale-up of INSTI-based regimens. However, the presence of accessory mutations calls for the surveillance of INSTI resistance mutations to ensure that the anticipated long-term desired outcomes are achieved.

The efficacy of immune checkpoint inhibitors in advanced EGFR-Mutated non-small cell lung cancer after resistance to EGFR-TKIs: Real-World evidence from a multicenter retrospective study.

BackgroundThe efficacy of immune checkpoint inhibitors (ICIs) in pretreated EGFR-mutated non-small cell lung cancer (NSCLC) patients is controversial. We conducted this multicenter retrospective study to examine the efficacy of ICIs in a real world setting.Patients and methodsWe collected 116 consecutive NSCLC patients with sensitive EGFR mutations who received ICIs alone or in combination after failure to respond to EGFR tyrosine kinase inhibitors (EGFR-TKIs), and 99 patients were included for final analysis. The impacts of ICIs on the patients’ objective response rate (ORR), disease control rate (DCR), progression-free survival (PFS), and overall survival (OS) were assessed. The relationships between outcomes and clinical characteristics were analyzed.ResultsThe ORR in patients with target lesions was 31.25% (95% CI: 22.18-41.52), and the DCR in all patients was 65.66% (95% CI: 55.44-74.91). The overall median PFS was 5.0 months (95% CI: 3.0-6.6), and the median OS was 15.9 months (95% CI: 10.8-23.8). The outcomes were better in patients receiving combination therapy with ECOG scores of 0-1 and no more than 2 lines of prior therapy, with a median PFS of 7.4 months (95% CI: 3.0-13.3) and a median OS of 29.0 months (95% CI: 11.7-NE). Primary EGFR mutation type and treatment mode were found to have a notable impact on clinical outcomes. Both median PFS and OS in patients with EGFR L858R mutation were significantly shorter than those in patients with EGFR exon 19 deletion (19del) (PFS: 2.5 versus 6.7 months, HR: 1.80, log-rank P=0.011; OS: 9.8 versus 26.9 months, HR: 2.48, log-rank P=0.002). Patients receiving combination therapy had notably longer median PFS and OS than those receiving monotherapy (PFS: 5.2 versus 3.0 months, HR: 0.54, log-rank P=0.020; OS: 19.0 versus 7.4 months, HR: 0.46, log-rank P=0.009).ConclusionsOur study suggests that ICI-based combination therapy is a potential strategy for EGFR-mutated NSCLC patients after EGFR-TKI failure. The efficacy may differ according to EGFR subtypes.

Genetic profile and characteristics of HIV-1 drug resistance mutation in the Krasnodar region over the 2014 to 2019

Objective.Molecular genetic analysis of AIDS-1, having spread in the territory of Krasnodar region over the 2014 to 2019, including analysis of drug resistance mutationMaterials and methods.The study used the collection of samples, obtained from 478 HIV-infected people, living in the territory of Krasnodar region over the 2014 to 2019; out of them 155 received antiretroviral therapy (ART). Genotyping of the obtained samples was carried out with the further phylogenetic analysis, including detailed analysis of identified unique recombinant forms of HIV-1.Results.It is shown that HIV-1 sub-subtype A6 (80,5%) continues dominating in the territory of Krasnodar region, genetic variant B (9,2%) is the second most common one. Also 11 unique recombinant forms (URFs) were discovered, 10 out of which are the result of A6 and B genetic variants recombination. Drug resistance mutations were registered with 114 patients, received ART (73,6%); primary mutations — M184V and A62V. Provisional indicator of transmissible drug resistance in the population of naïve patients is 4,6% (15/323); primary mutations — K103N и G190S.Conclusion.The analysis of genetic profile and HIV-1 drug resistance mutation is carried out in the territory of Krasnodar region over the 2014 to 2019.

Efficacy and Safety of Rezivertinib (BPI-7711) in Patients With Locally Advanced or Metastatic/Recurrent EGFR T790M-Mutated NSCLC: A Phase 2b Study

IntroductionRezivertinib (BPI-7711) is a novel third-generation EGFR tyrosine kinase inhibitor (TKI) targeting both EGFR-sensitizing mutations and EGFR T790M mutation. This study aimed to evaluate the efficacy and safety of rezivertinib in patients with locally advanced or metastatic/recurrent EGFR T790M-mutated NSCLC. MethodsPatients with locally advanced or metastatic/recurrent NSCLC with confirmed EGFR T790M mutation who progressed after first-/second-generation EGFR TKI therapy or primary EGFR T790M mutation were enrolled. Patients received rezivertinib at 180 mg orally once daily until disease progression, unacceptable toxicity, or withdrawal of consent. The primary end point was objective response rate (ORR) assessed by blinded independent central review per Response Evaluation Criteria in Solid Tumors version 1.1. Secondary end points included disease control rate (DCR), duration of response, progression-free survival (PFS), overall survival, and safety. This study is registered with Clinical Trials.gov (NCT03812809). ResultsA total of 226 patients were enrolled from July 5, 2019, to January 22, 2020. By the data cutoff date on January 24, 2022, the median duration of follow-up was 23.3 months (95% confidence interval [CI]: 22.8–24.0). The ORR by blinded independent central review was 64.6% (95% CI: 58.0%–70.8%), and DCR was 89.8% (95% CI: 85.1%–93.4%). The median duration of response was 12.5 months (95% CI: 10.0–13.9), and median PFS was 12.2 months (95% CI: 9.6–13.9). The median overall survival was 23.9 months (95% CI: 20.0–not calculated [NC]). Among 91 (40.3%) patients with central nervous system (CNS) metastases, the median CNS PFS was 16.6 months (95% CI: 11.1–NC). In 29 patients with more than or equal to one brain target lesion at baseline, the CNS ORR and CNS DCR were 69.0% (95% CI: 49.2%–84.7%) and 100% (95% CI: 88.1%–100%), respectively. Time to progression of CNS was 16.5 months (95% CI: 9.7–NC). Of 226 patients, 188 (83.2%) had at least one treatment-related adverse event, whereas grade more than or equal to 3 occurred in 45 (19.9%) patients. No interstitial lung disease was reported. ConclusionsRezivertinib was found to have promising efficacy and favorable safety profile for patients with locally advanced or metastatic/recurrent NSCLC with EGFR T790M mutation.

Compassionate use of long-acting cabotegravir plus rilpivirine for people living with HIV-1 in need of parenteral antiretroviral therapy.

Physicians could request compassionate use of oral and long-acting (LA) cabotegravir + rilpivirine for people living with HIV-1 under a single-patient request programme supported by ViiV Healthcare and Janssen. Outcomes are reported. Eligibility criteria included need for parenteral therapy, no primary resistance mutations to cabotegravir or rilpivirine, and established retention in care. Demographic, efficacy, and safety data were obtained from standardized programme applications and quarterly clinical updates. Individuals received a loading dose of LA cabotegravir 600 mg + rilpivirine 900 mg, followed by LA maintenance doses of 400 mg/600 mg every 4 weeks; some received lead-in oral cabotegravir and rilpivirine. Through July 2020, 35 people living with HIV-1 had data available. The most frequent reason for compassionate use request was chronic non-adherence due to psychological conditions (n=15). Of 35 people living with HIV-1, 28 had detectable viremia (median viral load 60 300 copies/mL) and seven were virologically suppressed at programme entry; 16/28 and 6/7 achieved or maintained virological suppression at data cutoff, respectively. Seven people living with HIV-1 discontinued for incomplete virological response, six with detectable viremia at initiation; six and four had new reverse transcriptase and integrase mutations at discontinuation, respectively. Six non-fatal serious adverse events were reported, two considered possibly treatment related. Four deaths were reported; none were treatment related. One individual reported two pregnancies and continued LA dosing. Most people living with HIV-1 had advanced disease and achieved (16/28) or maintained (6/7) virological suppression with LA therapy. Cabotegravir LA + rilpivirine LA as compassionate use provided a valuable treatment option for individuals with adherence issues with oral therapy and advanced disease.

KMT2C mutation in a Chinese man with primary multidrug-resistant metastatic adenocarcinoma of rete testis: a case report

BackgroundAdenocarcinoma of the rete testis (AORT) is an extremely rare malignant tumor with poor prognosis and limited responsiveness to traditional chemotherapy. Few previous studies have focused on the molecular mechanisms underlying therapy resistance in AORT and further scrutiny is required to enable searches for targeted drugs to guide treatment selection.Case presentationThe current case concerns a 55-year-old man with AORT who presented with isolated bone metastasis at initial diagnosis and experienced rapid disease progression after multi-line platinum-based combination chemotherapy. Next-generation sequencing revealed a novel somatic lysine methyltransferase 2C (KMT2C) c.5605 T > C mutation in exon 36 with an abundance of 49.27%. The patient received antiangiogenic drug treatment for 2 months but this was discontinued due to unacceptable anorexia and nausea. He survived for 12 months after diagnosis.ConclusionA potential correlation between AORT primary multi-drug resistance and KMT2C mutations is implied. Further studies are needed to determine the efficacy of PARP1/2 inhibitors for tumors with KMT2C mutations.

Traditional Chinese Medicine in Treating Primary Podocytosis: From Fundamental Science to Clinical Research.

Podocytes form a key component of the glomerular filtration barrier. Damage to podocytes is referred to as “podocyte disease.” There are many causes of podocyte injury, including primary injury, secondary injury, and gene mutations. Primary podocytosis mostly manifests as nephrotic syndrome. At present, first-line treatment is based on glucocorticoid administration combined with immunosuppressive therapy, but some patients still progress to end-stage renal disease. In Asia, especially in China, traditional Chinese medicine (TCM) still plays an important role in the treatment of kidney diseases. This study summarizes the potential mechanism of TCM and its active components in protecting podocytes, such as repairing podocyte injury, inhibiting podocyte proliferation, reducing podocyte apoptosis and excretion, maintaining podocyte skeleton structure, and upregulating podocyte-related protein expression. At the same time, the clinical efficacy of TCM in the treatment of primary podocytosis (including idiopathic membranous nephropathy, minimal change disease, and focal segmental glomerulosclerosis) is summarized to support the development of new treatment strategies for primary podocytosis.

Pyrimidine derivatives as EGFR tyrosine kinase inhibitors in non-small-cell lung cancer: A comprehensive review.

EGFR-positive non-small-cell lung cancer (NSCLC) due to primary mutation (EGFR DEL19 & L858R) has been recognized as a crucial mediator of tumor progression. This led to the development and approval of EGFR tyrosine kinase inhibitors, which addresses EGFR-mediated NSCLC but fail to show potency after initial months of therapy due to acquired resistance (EGFR T790M, EGFR C797S). Extensive research allowed identification of drugs for EGFR-positive NSCLC, wherein the majority of compounds have a pyrimidine substructure offering marked therapeutic benefits compared with chemotherapy. This current review outlines the diverse pyrimidine derivatives with amino-linked and fused pyrimidine scaffolds such as furo-pyrimidine, pyrimido-pyrimidine, thieno-pyrimidine, highlighting pyrimidine EGFR TK inhibitors reported in research emphasizing structural aspects, design approaches, inhibition potential, selectivity profile toward mutant EGFR conveyed through biological evaluation studies. Furthermore, mentioning the in-silico interaction profile of synthesized compounds for evaluating the binding affinity with key amino acids. The epilogue of review focuses on the recent research that drives forward to aid in the discovery and development of substituted amino and fused scaffolds of pyrimidine that can counteract the mutations and effectively manage EGFR-positive NSCLC.

TRAPS mutations in Tnfrsf1a decrease the responsiveness to TNFα via reduced cell surface expression of TNFR1.

Tumor necrosis factor (TNF) receptor-associated periodic syndrome (TRAPS) is an autoinflammatory periodic fever syndrome associated with heterozygous mutations in TNFRSF1A, which encodes TNF receptor type I (TNFR1). Although possible proinflammatory mechanisms have been proposed, most previous studies were performed using in vitro overexpression models, which could lead to undesirable inflammatory responses due to artificial overexpression. It is crucial to reproduce heterozygous mutations at physiological expression levels; however, such studies remain limited. In this study, we generated TRAPS mutant mice and analyzed their phenotypes. Three Tnfrsf1a mutant strains were generated by introducing T79M, G87V, or T90I mutation. T79M is a known mutation responsible for TRAPS, whereas G87V is a TRAPS mutation that we have reported, and T90I is a variant of unknown significance. Using these murine models, we investigated whether TRAPS mutations could affect the inflammatory responses in vivo and in vitro. We found that none of the mutant mice exhibited detectable inflammatory phenotypes under standard housing conditions for 1 year. Interestingly, TRAPS mutant (T79M and G87V) mice had reduced mortality rates after the administration of lipopolysaccharide (LPS) and D-galactosamine, which induce TNFα-dependent lethal hepatitis. Moreover, TRAPS mutations strongly suppressed the development of TNFα-mediated arthritis when crossed with human TNFα transgenic mice. In in vitro primary bone marrow-derived macrophage cultures, the T79M and G87V mutations attenuated the inflammatory responses to TNFα compared with the wild-type, whereas these mutations did not alter the responsiveness of these cells to LPS. The T90I mutant macrophages behaved similarly to wild type in response to LPS and TNFα. The TNFR1 levels were increased in whole-cell lysates of TRAPS mutant macrophages, whereas the cell surface expression of TNFR1 was significantly decreased in TRAPS mutant macrophages. Taken together, TRAPS mutations did not augment the inflammatory responses to TNFα and LPS; instead, they suppressed the response to TNFα via decreased cell surface expression of TNFR1. The stimulation of lymphotoxin-α, adenosine triphosphate, and norepinephrine in primary macrophages or various stimuli in murine splenocytes did not induce detectable inflammatory responses. In conclusion, TRAPS mutations suppressed responsiveness to TNFα, and TRAPS-associated inflammation is likely induced by unconfirmed disease-specific proinflammatory factors.

Discovery of BLU-945, a Reversible, Potent, and Wild-Type-Sparing Next-Generation EGFR Mutant Inhibitor for Treatment-Resistant Non-Small-Cell Lung Cancer.

While epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have changed the treatment landscape for EGFR mutant (L858R and ex19del)-driven non-small-cell lung cancer (NSCLC), most patients will eventually develop resistance to TKIs. In the case of first- and second-generation TKIs, up to 60% of patients will develop an EGFR T790M mutation, while third-generation irreversible TKIs, like osimertinib, lead to C797S as the primary on-target resistance mutation. The development of reversible inhibitors of these resistance mutants is often hampered by poor selectivity against wild-type EGFR, resulting in potentially dose-limiting toxicities and a sub-optimal profile for use in combinations. BLU-945 (compound 30) is a potent, reversible, wild-type-sparing inhibitor of EGFR+/T790M and EGFR+/T790M/C797S resistance mutants that maintains activity against the sensitizing mutations, especially L858R. Pre-clinical efficacy and safety studies supported progression of BLU-945 into clinical studies, and it is currently in phase 1/2 clinical trials for treatment-resistant EGFR-driven NSCLC.

High Prevalence of Hemophagocytic Lymphohistiocytosis in Acute Liver Failure of Infancy

To investigate the prevalence of hemophagocytic lymphohistiocytosis (HLH) syndrome in pediatric acute liver failure (PALF) of infancy and assess the diagnostic role of rapid immunologic tests, genotype/phenotype correlations, and clinical outcomes. We retrospectively analyzed 78 children with PALF aged <24months referred over almost 2 decades. The studied patients with a phenotype of HLH syndrome had a comprehensive immunologic workup, including additional genetic analysis for primary immunologic causes. Thirty of the 78 children had the HLH phenotype and underwent genetic assessment, which demonstrated positive findings in 19 (63.3%), including 9 (30%) with biallelic primary HLH mutations and 10 (33.3%) with heterozygous mutations and/or polymorphisms. The most common form of primary HLH was familial hemophagocytic lymphohistiocytosis (FHL)-2, diagnosed in 6 children, 4 of whom had a c.50delT (p.Leu17ArgfsTer34) mutation in the PRF1 gene. Three patients with primary HLH received genetic diagnoses of FHL-3, Griscelli syndrome, and LRBA (lipopolysaccharide-responsive vesicle trafficking, beach- and anchor-containing) protein deficiency. Overall mortality in the series was 52.6% (10 of 19), and mortality in children with a documented biallelic pathogenic HLH mutation (ie, primary HLH) was 66.6% (6 of 9). Two children underwent liver transplantation, and 4 children underwent emergency hematopoietic stem cell transplantation; all but 1 child survived medium term. Primary HLH can be diagnosed retrospectively in approximately one-third of infants with indeterminate PALF (iPALF) who meet the clinical criteria for HLH, often leading to their death. The most common HLH type in iPALF is FHL-2, caused by biallelic mutations in PRF-1. The clinical relevance of observed heterozygous mutations and variants of uncertain significance requires further investigation. Prompt hematopoietic stem cell transplantation could be life-saving in infants who survive the liver injury.

High-level dolutegravir resistance can emerge rapidly from few variants and spread by recombination: implications for integrase strand transfer inhibitor salvage therapy.

The integrase strand transfer inhibitor (INSTI) dolutegravir is commonly used in combination antiretroviral therapy regimens and retains strong potency even with primary resistance mutations to some other INSTIs. Acquisition of accessory mutations to primary mutations results in significant increases in dolutegravir resistance. Previously, we reported that addition of the secondary mutation T97A can result in rapid treatment failure in individuals with INSTI mutations at positions 140 and 148. Here, we conducted a detailed case study of one of these individuals and find that T97A-containing HIV emerged from a large replicating population from only a few (≤4) viral lineages. When combined with primary INSTI resistance mutations, T97A provides a strong selective advantage; the finding that T97A-containing variants spread by replication and recombination, and persisted for months after discontinuing dolutegravir, has important implications as dolutegravir is rolled out worldwide.

Audit of Molecular Mechanisms of Primary and Secondary Resistance to Various Generations of Tyrosine Kinase Inhibitors in Known Epidermal Growth Factor Receptor-Mutant Non-small Cell Lung Cancer Patients in a Tertiary Centre

AimsPresently, three generations of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are approved against oncogene addicted EGFR-mutant non-small cell lung cancer (NSCLC). Patients with actionable EGFR mutations invariably develop resistance. This resistance can be intrinsic (primary) or acquired (secondary). Materials and methodsThis was a retrospective study carried out between January 2016 and April 2021 analysing 486 samples of NSCLC for primary and secondary resistance to first- (erlotinib, gefitinb), second- (afatinib) and/or third-generation (osimertinib) TKIs in EGFR-mutant NSCLCs by next generation sequencing (NGS). Tissue NGS was carried out using the Thermofischer Ion Torrent™ Oncomine™ Focus 52 gene assay; liquid biopsy NGS was carried out using the Oncomine Lung Cell-Free Total Nucleic Acid assay. All cases were previously tested for a single EGFR gene with the Therascreen® EGFR RGQ PCR kit. ResultsThe results were divided into four groups: (i) group 1: primary resistance to first- and/or second-generation TKIs. This group, with 21 cases, showed EGFR exon 20 insertions, dual, complex mutations and variant of unknown significance, de novo MET gene amplification besides other mutations. (ii) Group 2: primary resistance to third-generation TKIs. This group showed two cases, with one showing dual EGFR mutation (L858R and E709A) and EGFR gene amplification. (iii) Group 3: secondary resistance to first- and second-generation TKIs. This group had 27 cases, which were previously reported negative for EGFR T790M by single gene testing. Significant findings were MET gene amplification in four cases, with one also showing MET exon 14 skipping mutation. Three cases showed small cell change and one showed loss of primary mutation. (iv) Group 4: secondary resistance to third-generation TKIs. The latter group was further subgrouped into group 4A: secondary resistance to osimertinib (third-generation TKI) when offered as second-line therapy after first- and second-generation TKIs on detection of T790M mutation. This group had 15 cases. EGFR T790M mutation was lost in 10 (10/15; 67%) cases and was retained in five cases. Patients with T790M loss experienced early resistance (6.9 months versus 12.6 months mean, P = 0.0024) compared with cases that retained T790M. Two cases gained MET amplification as the resistance mechanisms. Other mutations that were found when EGFR T790M was lost were in FGFR3, KRAS, PIK3CA, CTNNB1, BRAF genes. One case had EML4-ALK translocation. Two cases showed driver EGFR deletion 19, retained T790M and C797S mutation in Cis form. Group 4B: secondary resistance to osimertinib (when given as first-line therapy) in EGFR-mutant NSCLC. This group had three cases. The duration of osimertinib treatment ranged from 11 to 17 months. Two patients showed additional C797S mutation along with primary EGFR mutation. ConclusionThis study shows the wide spectrum of primary and secondary EGFR resistance mechanisms to first, second and third generation of TKIs and helps us to identify newer therapeutic targets that could carry forward the initial advantage offered by EGFR TKIs.

Susceptibility of Drug Resistant Hepatitis B Virus Mutants to Besifovir.

Currently, interferon alpha and nucleos(t)ide analogues (NAs) are clinically available to treat hepatitis B virus (HBV) infection. Several NAs, including lamivudine (LMV), adefovir (ADV), entecavir (ETV) and tenofovir (TDF or TAF) have been approved and administered to chronic hepatitis B (CHB) patients. NAs inhibit HBV DNA synthesis by targeting the reverse transcriptase (RT) domain of HBV polymerase. Several mutations in the RT domain which lead to drug resistance against NAs have been reported, even for TDF and TAF which are highly potent with very low resistance rate. Besifovir (BFV) is a new antiviral dGMP analogue able to be used as a new NA drug for the control of CHB infection. Drug resistance to BFV is not well known due to its shorter duration of clinical use. Recently, we reported that rtL180M (M) and rtM204V (V) mutations, already resistant to LMV, are associated with BFV resistance. However, the susceptibility to BFV of previously known HBV mutants resistant to various drugs has not been studied. To investigate this, we performed in vitro drug susceptibility assays using natural and artificial mutants that are associated with resistance to LMV, ADV, ETV or TDF. As a result, LMV-resistant mutants were not susceptible to BFV and ETV-resistant clones showed partial resistance against BFV as well. However, ADV-resistant mutants were highly sensitive to BFV. In case of tenofovir-resistant mutations, the HBV mutants harboring primary mutations to tenofovir resistance were susceptible to BFV. Therefore, our study revealed that BSV may serve as an alternative drug for patients with ADV-, ETV-, TDF- or TAF-resistance.

Efficacy and Safety of SH-1028 in Patients With EGFR T790M-Positive NSCLC: A Multicenter, Single-Arm, Open-Label, Phase 2 Trial

IntroductionAs a novel third-generation EGFR tyrosine kinase inhibitor (TKI), SH-1028 (formerly oritinib) is developed to inhibit both sensitizing EGFR mutations and EGFR T790M mutation. MethodsThis was a multicenter, single-arm, open-label, phase 2 trial (NCT03823807). Eligible patients were those with advanced NSCLC with centrally confirmed EGFR T790M mutation who progressed after first- or second-generation EGFR TKIs or with primary EGFR T790M mutations. Each patient received SH-1028 tablets orally at 200 mg/d until disease progression or intolerable toxicity. Tumor response was evaluated every 6 weeks per the Response Evaluation Criteria in Solid Tumors, version 1.1. The primary end point was objective response rate by an independent review committee. The secondary end points were progression-free survival, overall survival (OS), disease control rate, safety, and so on. ResultsA total of 286 patients with EGFR T790M-positive advanced NSCLC were enrolled in this study, including 59 patients in part A (dose-verification study) and 227 patients in part B (second-line registration study). By data cutoff on September 17, 2021, the independent review committee–assessed objective response rate was 55.9% (95% confidence interval [CI]: 42.4–68.8) in part A and 60.4% (95% CI: 53.7–66.8) in part B. The median progression-free survival was 12.4 months (95% CI: 8.3–20.8) in part A and 12.6 months (95% CI: 9.7–15.3) in part B. The median OS was 26.0 months (95% CI: 23.3–not reached) in part A, and OS was immature in part B. Among the 286 patients, 44 of them experienced at least one grade 3 or higher treatment-related adverse event, with the most common ones as increased serum creatinine phosphokinase level (13 [4.5%]), diarrhea (six [2.1%]), and prolonged QT interval (three [1.0%]). Treatment-related skin rash was reported in 26 patients (9.1%), all grade 1 or 2. There was no interstitial lung disease reported in this study. ConclusionsSH-1028 is efficacious and tolerable in second-line treatment of patients with advanced NSCLC with positive EGFR T790M.

Analysis of the entire mitochondrial genome reveals Leber’s hereditary optic neuropathy mitochondrial DNA mutations in an Arab cohort with multiple sclerosis

Several mitochondrial DNA (mtDNA) mutations of Leber's hereditary optic neuropathy (LHON) have been reported in patients with multiple sclerosis (MS) from different ethnicities. To further study the involvement of LHON mtDNA mutations in MS in the Arab population, we analyzed sequencing data of the entire mitochondrial genome from 47 unrelated Saudi individuals, 23 patients with relapse-remitting MS (RRMS) and 24 healthy controls. Ten LHON mutations/variants were detected in the patients but were absent in the controls. Of them, the common primary pathogenic mutation m.14484T>C and the rare mutation m.10237T>C were found in one patient, whereas the rare mutation m.9101T>C was found in another patient. The remaining were secondary single nucleotide variants (SNVs) found either in synergy with the primary/rare mutations or individually in other patients. Patients carrying LHON variants also exhibited distinct mtDNA variants throughout the mitochondrial genome, eight were previously reported in patients with LHON. Moreover, five other LHON-related SNVs differed significantly in their prevalence among patients and controls (P < 0.05). This study, the first to investigate LHON mtDNA mutations/variants in a Saudi cohort may suggest a role of these mutations/variants in the pathogenesis or genetic predisposition to MS, a possibility which needs to be explored further in a large-scale.

Genetic features of TP53 mutation and its downstream FOXA1 in prostate cancer.

Metastasis is the most lethal form of prostate cancer, and finding new therapeutic targets remains a major clinical challenge. TP53 mutation has been identified to be involved in tumor progression and metastasis. Nevertheless, direct evidence of the role of TP53 mutation in prostate cancer metastasis and its underlying mechanism remain obscure. Herein, TP53 was found to be the most mutated gene in prostate cancer, and missense mutations were the primary mutation type based on bioinformatics data analysis. Subsequently, TP53 rs12947788 mutation site was significant in prostate cancer, and correlated with metastasis and tumor-node-metastasis (TNM) stage. Furthermore, forkhead box A1 (FOXA1), a target of TP53, was highly expressed in prostate cancer tissue, especially in TP53-mutant patients. It was also associated with patients' Gleason scores and nodal metastasis. Knockdown of FOXA1 suppressed the migration in prostate cancer cells in vitro. Our findings indicate that targeting TP53 mutation and FOXA1 might be a promising therapeutic target for prostate cancer metastasis.

Molecular heterogeneity and analysis of the long-term survival of patients with gastrointestinal stromal tumors

Introduction. Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract the character diagnostic feature of which is CD117 (KIT) expression. GISTs are clinically diverse and have different genetic alterations that may have predictive and prognostic significance.Aim – the study of clinical, morphological and genetic features of GISTs to assess the overall survival (OS) of patients with various profiles of genetic disorders for elucidation the factors contributing to prognosis.Materials and methods. A total 244 GIST patients who received combined treatment were enrolled in the study and their clinical characteristics and mutational status of KIT, PDGFRA, BRAF were analyzed. SDH-deficient GISTs were detected using IHC-analysis of SDHB expression.Results. Stromal tumors developed in stomach (50 %), small intestine (37.7 %), colon or rectum (8.6 %), esophagus (0.4 %) and extraorganically (EGIST, 5.7 %). Overall survival correlated with gastric site (p = 0.005), tumor size &lt;10 cm (p = 0,0001) and mitotic count HPF&lt; 10 / 50 (p = 0.007). KIT mutations were found in 168 (68.9 %) and PDGFRA – in 31 (12.1 %) of GISTs, 14 novel mutations were detected. Mutations in KIT exon 11 were found in 140 (57.4 %) tumors, 10-year OS, 51 %, median 124 months. Patients with deletions had lower OS than patients with substitutions or duplications in KIT exon 11 (p = 0,023). The lowest OS was in patients with primary mutations in KIT exons 13 or 17 (median 28 months) and duplications in KIT exon 9 (median 71 months). There was a low OS of young patients with homozygous KIT mutations, mutations that begin in intron and two simultaneous KIT mutations. GISTs with PDGFRA mutations were located in stomach and had no metastases, 10-year OS, 63 %, median 175 months. KIT / PDGFRA mutations were not observed in 45 (18.4 %) patients (wild-type GIST), 10-year OS, 59 %, median 250 months. Wild-type GISTs with BRAF, NF1 mutations and SDH deficiency were detected. The better OS was demonstrated by patients with BRAFV600E (10-year ОS, 84 %, median 97 months) and SDH deficiency (10-year and 15-year OS, 82 %).Conclusion. Genetic analysis is necessary to clarify GIST prognosis and predict the effectiveness of targeted therapy. The clinical, morphological and genetic diversity of GISTs was confirmed. Wild-type GISTs with BRAF mutations and SDHdeficiency were identified in the Russian population for the first time. The long-term 10- и 15-year OS of GIST patients were evaluated.