Del Mutation Research Articles

Genetic analysis of a novel FBN1 mutation in a pediatric Marfan syndrome patient.

The aim of this study was to investigate a novel FBN1 gene mutation in a pediatric patient with Marfan syndrome (MFS) to provide a theoretical basis for genetic counseling. The subject was a 5-month-old male infant. With informed consent from the proband and his family, 2 mL of peripheral venous blood was collected from the patient, his father, mother, and sister. DNA was extracted using a DNA extraction kit with EDTA-K as an anticoagulant. The extracted DNA was subjected to minigene transcription and bioinformatics analysis. For minigene construction, wild-type and mutant minigenes were inserted into pcMINI and pcMINI-C vectors, respectively. Four recombinant vectors were transfected into the HeLa and 293T cell lines. After transfection for 48 hours, RNA was extracted from eight samples. DNA was also extracted from the family members' samples to construct a library. Target regions were captured using the SureSelect Human All Exon V6 (Agilent) kit and were sequenced with Illumina NovaSeq (sequencing read length 2×150 bp). Bioinformatic analysis identified the c.8226+5del mutation as a variant of uncertain clinical significance (VOUS). Literature and database reviews confirmed that this mutation had not been previously reported, identifying it as a novel mutation. The study identified a novel FBN1 mutation, c.8226+5del, that may be associated with clinical features such as low-set ears and distinctive facial characteristics in the proband. This mutation likely affects normal mRNA splicing, altering the structure and function of Exon 64 and potentially contributing to the development of autosomal dominant MFS.

The incidence of TP53 mutations in patients with chronic lymphocytic leukemia in Iran

Objectives Chronic lymphocytic leukemia (CLL) is linked to a highly variable disease course regarding responses to chemoimmunotherapy and clinical outcomes. Mutations in TP53 and/or deletions in chromosome 17p locus [del(17p)] may lead to loss of a TP53 allele. We investigated TP53 mutations in patients with CLL. Material and Methods Thirty patients with CLL, aged 40–84 years, were included. Immunophenotyping of B cells was done using flow cytometry by CD5, CD10, CD19, CD20, CD23, CD49d, CD38, FMC7, and CD200 markers. Bone marrow aspiration and peripheral blood smear examination were also performed. DNA was extracted and sequencing was done using the Sanger method. Structural aberrations were investigated using the FISH method. Results TP53 mutations had a frequency of eight cases (23.3%), three of which were missense mutations (37.5%), three were intronic mutations (37.5%), and two were silent mutations (25%). Peripheral B lymphocytes had a mean of 82% with 4.59% prolymphocytes. Conclusion Accurate testing for TP53 mutations [TP53 and del(17p) mutations] before every treatment line enables us to make proper therapeutic decisions to improve patient outcomes.

Identification and functional characterization of compound heterozygous CYP11B1 gene mutations.

11β-Hydroxylase deficiency (11β-OHD) is the second leading cause of congenital adrenal hyperplasia (CAH), a rare autosomal recessive disease caused by mutations in the CYP11B1 gene. We previously reported the case of a male Chinese patient with typical 11β-OHD symptoms. Sanger sequencing revealed that the patient carried a splice-site mutation, c.595+1G>A in the CYP11B1 gene. His mother and sister harbored the heterozygous mutation, c.595+1G>A. Paradoxically, Sanger sequencing did not detect any abnormality in the CYP11B1 gene of his father and brother. Therefore, in this study, we aimed to further explore the exact genetic etiology of 11β-OHD in this pedigree and analyze the functional consequence of the c.595+1G>A mutation. Gemomic DNA was extracted from the peripheral blood leukocytes of the family members and normal control individuals, followed by quantitative real-time polymerase chain reaction (qPCR) to detect the copy number of the target CYP11B1 gene fragment. Mutation analysis was also performed via whole-exome sequencing (WES) followed by Sanger sequencing validation. In vitro minigene assay was also performed to investigate the impact of the c.595+1G>A mutation on pre-mRNA splicing. qPCR results suggested a heterozygous deletion encompassing position c.595+1 along with flanking exonic and intronic sequences in the CYP11B1 gene of the patient and his father. WES followed by Sanger sequencing verified that the patient carried compound heterozygous mutations in the CYP11B1 gene, including a novel 2840-bp deletion (c.395+661_c.1121+180del) and c.595+1G>A, while his father carried the heterozygous c.395+661_c.1121+180del mutation. No other novel CYP11B1 mutations were found in the rest of the family members. Furthermore, minigene assay revealed that the c.595+1G>A mutation resulted in a 70-bp deletion of exon 3 in the mRNA, and this altered the reading frame at amino acid 176 and created a premature stop codon at amino acid 197. We identified a novel 2840-bp-sized large deletion and confirmed that the c.595+1G>A mutation disrupts normal pre-mRNA splicing. Either mutation could significantly alter the reading frame and abolish CYP11B1 enzyme activity. Therefore, our findings widen the mutation spectrum of CYP11B1 and provide an accurate diagnosis of 11β-OHD at a molecular genetic level.

Analyzing three pedigrees in X-linked Alport syndrome with the presentation of nephrotic syndrome.

Alport syndrome (AS) is a common cause of end-stage renal disease (ESRD) with various clinical symptoms and incomplete manifestation. Patients with AS and other renal disorders are often misdiagnosed. This study reported three X-linked dominant Alport syndrome (XLAS) pedigrees with nephrotic syndrome (NS) as the predominant phenotype and analyzed COL4A5 gene alterations. Three Han Chinese XLAS pedigrees were recruited, and clinical phenotypes were obtained. The pre-certified individuals' peripheral blood DNA was taken, and whole-genome next-generation sequencing (NGS) was performed for candidate genes and mutation screening, followed by NGS or Sanger sequencing of suspected mutant types in participating family members. Both probands A and B were diagnosed with NS through biochemical tests, and X-linked Alport syndrome-associated renal injury was diagnosed by renal biopsy. The biopsy revealed focal foamy cells in the renal interstitium, tearing and delamination changes in the glomerular basement membrane, and negative α3 and α5 chains of type IV collagen. Proband C, who was earlier diagnosed with NS, has now advanced to ESRD, along with his mother and proband A's mother. Genetic sequencing of all three pedigrees identified three mutations, namely, c.5020C>T, c.4435_4445del, and c.1584_1587+6del in the X-linked dominant gene COL4A5 (NM_000495.5). These mutations lead to the production of shortened proteins, potentially impacting the function of COL4A5 and causing pathogenic effects. The novel c.4435_4445del and c.1584_1587+6del mutations not only enrich the spectrum of mutations in the COL4A5 gene but also indicate that carriers of both mutation sites and those with mutation c.5020C>T may present NS as their primary clinical manifestation.

Real-World Outcomes Among 2L+ Patients with Chronic Lymphocytic Leukemia (CLL) By Line of Therapy (LOT)

Introduction: The chronic lymphocytic leukemia (CLL) treatment landscape is rapidly evolving, and real-world (rw) outcomes of patients receiving 2L+ therapy in the current treatment paradigm are incompletely understood. This study examined the rw outcomes of patients with CLL/ small lymphocytic lymphoma (SLL) receiving 2 or more lines of therapy (LOTs) using a large, contemporary rw dataset. Methods: Patients meeting the study criteria were identified in the COTA real-world, clinical database: Aged ≥ 18 years at diagnosis with CLL/SLL who initiated 2L therapy between January 1, 2014 and June 30, 2021. The initial index date for the study was the date of 2L initiation and was updated to the start of each subsequent LOT for LOT-specific analyses. The following time to event outcomes were evaluated for the study population and by LOT using the Kaplan-Meier method: time to next treatment (TTNT: time from index to the initiation of the next LOT or death), time to treatment discontinuation (TTD: time from index to treatment discontinuation for any reason, including end of therapy as prescribed/indicated), rw duration of response (rwDOR: time from the first clinician-documented CR or PR to the first assessment of progressive disease or the initiation of next LOT among responders), rw progression-free survival (rwPFS), and rw overall survival (rwOS). For all time-to-event endpoints, patients were censored at the date of last visit. Rw overall response rate (rwORR) was the proportion of patients who had a clinician-documented PR or CR as the best response to a LOT among all patients who received that LOT. Results: A population of 1,102 patients was identified from the COTA database meeting the study criteria. Key patient and clinical characteristics included: median age at diagnosis of 64 years, 60.9% male, 88.1% treated in a community setting, 74.1% diagnosed in 2014 or earlier, 9.3% positive for TP53 mutation (55.0% unknown), and 7.8% with del(17p) mutation (66.2% unknown). Median TTNT among patients who received 2L (n=1102) was 29.3 months (95% CI: 27.3, 32.6), and median DOR to 2L (n=863) was 31.9 months (95% CI: 27.5, 36.4). Generally, median TTNT and DOR decreased across LOT 2-4. Median TTNT from 3L (n=468) and 4L (n=201) initiation were 21.4 (95% CI: 18.2, 25.1) and 15.1 months (95% CI: 11.5, 20.2), respectively, and median DOR among 3L (n=333) and 4L (n=130) responders were 22.6 (95% CI: 18.3, 26.2) and 15.1 months (95% CI: 11.7, 22.9), respectively. More than three-quarters (78.3%) of 2L patients experienced a physician-documented CR or PR, and rwORR decreased over subsequent LOTs (71.2% in 3L and 64.7% in 4L). Median TTD varied by treatment type. Patients who received continuous therapy with Bruton's Tyrosine Kinase inhibitors (BTKis) experienced longer median TTD, while patients who received fixed-duration therapies experienced shorter median TTD. Median TTD among patients who received ibrutinib (n=391), acalabrutinib (n=54), or venetoclax + rituximab (n=48) in 2L was 24.8, 32.6, or 23.5 months, respectively, while median TTD among patients who received BR (n=194) or rituximab monotherapy (n=67) was 4.0 or 0.9 months, respectively. Finally, median rwPFS and rwOS decreased by LOT for patients who received 2L-4L. Median rwPFS for 2L, 3L, and 4L were 31.4 (95% CI: 28.6, 35.5), 22.8 (95% CI: 19.1, 26.4), and 16.5 (95% CI: 13.6, 22.6) months, respectively, and median rwOS were 79.0 (95% CI: 68.9, 85.3), 56.3 (95% CI: 51.6, 64.5), and 51.9 months (95% CI: 33.8, Not Reached), respectively. Conclusions: In this real-world, contemporary database analysis, median TTNT and DOR exceeded 2 years among 2L patients. Although the adoption of novel therapies has changed the CLL/SLL treatment paradigm and patients who received 2L BTKi therapy experienced numerically greater TTD relative to those who received other treatment types, patients with relapsed/refractory (R/R) disease continue to experience poor rwPFS and rwOS relative to patients in clinical trials. This study demonstrates the continued unmet need for this rw patient population and highlights the need for continued innovation, namely new therapeutic agents leveraging novel mechanisms of action, to improve rw outcomes for patients with R/R CLL/SLL.

Update on diagnosis and treatment of Chronic Lymphocytic Leukemia (CLL)

commonest leukemia found in adults western countries, affects more than 200000 people and is associated with approximately 4410 death in the USA annually. CLL is characterized by a progressive proliferation and accumulation of mature yet functionally incompetent lymphocytes. The disease typically occurs in elderly patients and has a highly variable clinical course. Leukemic transformation is initiated by specific genomic alterations that impair apoptosis of clonal B-cells. Diagnosis: The diagnosis of chronic lymphocytic leukemia (CLL) requires the presence of more than or equal to 5 × 109/L B lymphocytes (5000/μL) in the peripheral blood for the duration of at least 3 months. Immunophenotyping of circulating B-lymphocytes, which identify a clonal B-cell population carrying the CD5 antigen, as well as typical B-cell markers are confirmatory test. Prognosis: The two similar clinical staging systems, Binet and Rai stages, create prognostic information by using results of physical examination and blood counts. Various molecular biology markers also have prognostic value. Mutations of the TP53 gene and x chromosome 17 chromosome (del [17p) predict resistance to chemoimmunotherapy and a shorter time to progression, with most targeted therapies. A comprehensive, international prognostic score (CLL-IPI) integrates genetic, biological and clinical variables to identify distinct risk groups of CLL patients. Therapy: Early-stage disease of CLL patients are followed up regularly, only patients with active or symptomatic disease or with advanced stages of Binet or Rai require treatment. When treatment is indicated, most patients with CLL have several options: a combination of obinutuzumab and venetoclax, ibrutinib monotherapy, or immunochemotherapy. In patients under 65 years of age who are physically fit (particularly if they have a mutated IGVH gene), immunochemotherapy with fludarabine, cyclophosphamide, and rituximab remains the standard of care because of its potential therapeutic potential. In case of relapse, the initial treatment can be repeated if the treatment-free period exceeds 3 years. And in case of early recurrence of the disease, treatment should be switched to an alternative regime. Patients with del(17p) or TP53 mutations are another high-risk group and should be treated with targeted drugs. Allogeneic SCT can be considered in patients with TP53 or del(17p) mutations or in patients on inhibitor therapy.

EPS6.08 Cystic fibrosis elexacaftor/tezacaftor/ivacaftor in liver or kidney transplanted people with cystic fibrosis using Tacrolimus, a drug-drug interaction study

Background: Although elexacaftor/tezacaftor/ivacaftor (ETI) has a proven robust effect in people with cystic fibrosis (pwCF) with at least one F508 del mutation, its use is not recommended for pwCF after solid organ transplantation because of expected drug-drug interactions with antirejection therapy, mostly tacrolimus. With our drug-drug interaction study we aim to improve insight into the safety and clinical benefits of co-administration of ETI and tacrolimus in liver or kidney transplanted adult pwCF.

Survival Analysis of Lung Adenocarcinoma Patients with Exon 19 Del and 21 L858R Mutations Receiving EGFR-TKI Treatment

Introduction: Patients with adenocarcinoma of the lung that have a common EGFR mutation, the Exon 19 Del mutation, survive better than those with the Exon 21 L858R mutation. This study examined whether there is a significant difference in prognosis between two common EGFR mutations, namely exon 19 Del and 21 L858R. This study compared OS (overall survival) and PFS (progression-free survival) in NSCLC patients with Exon 19 Del and Exon 21 L858R mutations who received EGFR-TKI targeted therapy at H. Adam Malik Hospital Medan. Methods: This analysis study used a retrospective cohort design to evaluate the OS and PFS of NSCLC patients who underwent EGFR-TKI precision medicine at H. Adam Hospital Malik Medan between January 1, 2017, and December 31, 2020 and also had Exon 19 Del and Exon 21 L858R alterations. Results: A total of 88 people were sampled. The majority of research subjects were male (60.2%). Median OS was eleven months (95 percent CI:9.594-12,406). According to the study's data, eight people (9.1%) survived until the study's ending. The median OS of Exon 19 Del Common Mutation was 11 months (95%CI 9,064-12,936). While Exon 21 L858R group had ten months (95%CI 4,546-15,454). The log-rank test identified no statistical difference in median OS between mutation types (p=0.562). Conclusion: The findings of this study revealed that subjects with Exon 19 Del mutations had a longer median OS and PFS than those with Exon 21 L858R variants. Nevertheless, there was no significant difference in median OS and PFS between study subjects with mutation of Exon 19 Del and Exon 21 L858R, which received the targeted medication.

MET overexpression in EGFR L858R mutant treatment-naïve advanced lung adenocarcinoma correlated with poor prognosis: a real-world retrospective study.

Mesenchymal epithelial transition (MET) overexpression has been reported in approximately 50-60% of epidermal growth factor receptor (EGFR)-mutant non-small-cell lung cancers. However, the prognostic significance of MET overexpression has not been established in advanced lung adenocarcinoma (ADC) patients with EGFR-sensitive mutations. A retrospective study was performed on a total of 406 treatment-naïve advanced ADC patients with EGFR mutation detection and MET expression information. EGFR mutations were detected by next-generation sequencing or amplification refractory mutation system-polymerase chain reaction. Immuno-histochemistry staining of MET expression was evaluated by H-score and overexpression was defined as an H-score ≥ 200. Overall survival (OS) and progression-free survival (PFS) were analyzed according to MET expression. Among the 406 patients, 208 patients had EGFR mutations, including 102 exon 19_del mutations, 94 L858R mutations and 12 other types of mutations. Of 110 patients with concomitant EGFR mutations and MET overexpression, 61 (59.8%) patients had 19_del mutations, 44 (46.8%) patients had L858R mutations and five (41.7%) patients had others. Patients with MET overexpression had a markedly shorter PFS and OS than patients with MET H-score < 200 in the EGFR L858R mutation subgroup (median PFS: 12 versus 26months, p = 0.001; median OS: 24 versus 32months, p = 0.038), whereas no significant difference was observed in 19_del mutation subgroup. Multivariate Cox analysis showed that MET overexpression was an independent poor prognostic factor for PFS and OS in patients with the L858R mutations (HR = 3.064, 95% CI 1.705-5.507, p < 0.001; HR = 2.043, 95% CI 1.000-4.172; p = 0.049), rather than 19_del. MET overexpression is a poor prognostic factor for advanced ADC patients with the EGFR L858R mutation.

P783: SYSTEMIC THERAPY UTILIZATION AND HEMATOLOGIC OUTCOMES IN LOWER-RISK MYELODYSPLASTIC SYNDROMES (LR-MDS): FINDINGS FROM A REAL-WORLD MEDICAL RECORD REVIEW STUDY IN THE US, UK, AND EUROPE (EU)

Background: Patients (pts) with LR-MDS experience a mean life-year loss of 6 years, and the treatment focus is improvement of quality of life and prolongation of life expectancy. Aims: This retrospective medical record review assessed real-world data to determine prevailing LR-MDS treatment patterns and hematologic outcomes. Methods: Eligible pts (≥18 years of age) with LR-MDS (Jul 1, 2013–Sep 30, 2018) were identified by participating hematologist-oncologists in the US, Canada (CAN), UK, and EU (France, Germany, Spain). Study measures and hematologic outcomes were descriptively assessed (data abstraction Jun–Nov 2021). Aggregated analyses used pooled US/CAN and UK/EU data sets for the overall population and pt subgroups treated with clinician-defined first-line (1L) and second-line (2L) erythropoiesis-stimulating agents (ESAs). Results: Medical record data were abstracted by 114 US/CAN (n=174 pts; median age 60.6 years; 68.4% male) and 166 UK/EU clinicians (n=319 pts; median age 67.1 years; 72.4% male). Median follow-up duration from MDS diagnosis was 60.6 months (US/CAN) and 67.6 months (UK/EU). Approximately half of the pts had “low” risk status (49.4% US/CAN; 53.9% UK/EU) categorized by the Revised International Prognostic Scoring System at initial diagnosis. The most common karyotype abnormality was del(5q) (27.6% US/CAN; 20.4% UK/EU). SF3B1 mutation was recorded for 7.5% US/CAN and 8.2% UK/EU pts. Almost all pts had received ≥1 line of systemic treatment for managing MDS-associated anemia (US/CAN: 165 [94.8%] 1L, 77 [44.3%] 2L; UK/EU: 294 [92.2%] 1L, 133 [41.7%] 2L). In 1L, ESA-based regimens were the most common treatment with median duration of therapy of ≥2 years (US/CAN: 142 [86.1%], of which the majority (80.3%) was ESA monotherapy; UK/EU: 264 [89.8%], comprising 82.2% ESA monotherapy). Treatment characteristics and outcomes are shown in the Table. Among pts who were transfusion dependent (TD) before 1L ESA, a very small proportion achieved transfusion independence (TI) or red blood cell transfusion reduction ≥50%, which was maintained for a median of <1.5 years. Among the 1L ESA-treated pts who were TD, TI was achieved by 5.7% (US/CAN) and 12.9% (UK/EU). In 2L ESA-treated pts (n=23 US/CAN; n=46 UK/EU), almost all were previously treated with a 1L ESA-containing regimen (95.7% US/CAN; 84.8% UK/EU). The most common reasons for reinitiating ESA in 2L (n=22, 15.5% US/CAN; n=39, 14.8% UK/EU) were overall health status (40.9% US/CAN; 33.3% UK/EU), treatment efficacy (40.9% US/CAN; 48.7% UK/EU), and compliance with national guidelines (13.6% US/CAN; 46.2% UK/EU). Among the pts who reinitiated ESA in 2L, the most common reasons for discontinuation of 1L ESA were completion of planned course of therapy (45.5% US/CAN; 28.2% UK/EU), pt decision (40.9% US/CAN; 20.5% UK/EU), progressive disease (36.4% US/CAN; 51.3% UK/EU), and adverse events (13.4% US/CAN; 15.4% UK/EU). Lenalidomide was used in 1L therapy among 8.5% pts in US/CAN and 5.8% in UK/EU. Among those with del(5q) mutation, ESA was used in 1L therapy among 83.3% pts in US/CAN and 87.7% in UK/EU. Image:Summary/Conclusion: ESA-based regimens were the most common treatment for anemia management in pts with LR-MDS; >1/3 of pts were TD before initiating 1L ESA therapy. A considerable proportion of 1L ESA-treated pts reinitiated ESA in 2L despite the majority having discontinued 1L treatment due to planned therapy completion, pt decision, or progressive disease. This real-world study suggests that there is an unmet need in addressing anemia burden with ESAs, and novel and effective treatments are needed.

P814: CLINICAL CHARACTERISTICS AND GENE MUTATION ANALYSIS OF 148 CHILDREN WITH FANCONI ANEMIA IN CHINA

Background: Fanconi anemia (FA) is a rare autosomal recessive, X-linked (FANCB) or autosomal dominant (FANCR/RAD51) disease of bone marrow failure. Pathogenesis is mainly caused by gene mutation related to the FA pathway. At present, at least 23 genes have been discovered to play a role in the FA pathway. The gene mutations of FA have been reported in more and more countries. In China, there no large-scale cases have been reported. Aims: In this study, we analyzed the clinical characteristics of 148 FA children from 20 provinces in China from October 2003 to October 2021. We explored the relationship between genotype and phenotype in 105 children with FA genotyping results. This is the largest series of subtyped Chinese Fanconi anemia patients to date, and the results will be helpful for future clinical management. Methods: A total of 148 Fanconi anemia patients from 140 families were diagnosed by clinical phenotype, family history, chromosome breakage test induced by mitomycin C, Single cell gel electrophoresis experiment, targeted - sequence, whole exon sequencing, and MLPA method from October 2003 to October 2021. Since there were no genetic test results for the children before 2010, only the 105 patients after 2010 had gene sequencing results, which allowed genotyping. DNA damage repair defects were detected by MMC-induced Single cell gel electrophoresis and chromosome breakage experiments. Targeted-seq was used to identify the FA mutations of the patients. Multiplex ligation-dependent probe amplification (MLPA) was performed to detect large fragment deletions in patients with negative results in the targeted-seq but conforming to the FA by MMC-induced chromosome breakage experiments test and clinical manifestations. We performed WES of DNA samples from five patients and their parents. Results: 1) The most common subtype of FA in China is FA-A, followed by FA-D2 and FA-P。2) The most common deformities in FA-A patients are finger deformities and skin pigmentation, while heart deformities are more common in FA-D2/B/G/I/P subtypes. 3) The common mutations of FANCA are exon23c.2101A>G, exon28c.2778 + 1G>A, exon34c.3348 + 1G>A, exon21c.1844delC, and exon30 C2941T>G, which are different from Korea and Japan. 4) Homozygous nonsense mutation of exon32 c.3188G>A and exon29 c.2851C>T in FA-A likely benign, but the homozygous splicing mutation in exon29c.2852 + 1G>T is harmful. 4) Children with splicing and Del mutations of FANCD1 gene had poor clinical prognosis, while children with homozygous mutations of exon10c. 1792A>G is likely benign. 5) Cases of initial diagnosis of MDS or poor disease progression tend to occur in FA-B/C/E/G/J/L/M and FA-S subtypes. 6)Chromosome 1, 3, 7, and 8 abnormalities and mutations of SF3B1 P53 are related to disease progression. Summary/Conclusion: We made a comprehensive description and analysis of the clinical characteristics and mutant genes of Children with FA in China. We proposed the mutation sites related to clinical prognosis and the cytogenetic abnormalities of disease progression, providing new data for our comprehensive understanding of FA. Our data will be useful for FA future menagement.

Pertactin-Deficient Bordetella pertussis with Unusual Mechanism of Pertactin Disruption, Spain, 1986–2018

Bordetella pertussis not expressing pertactin has increased in countries using acellular pertussis vaccines (ACV). The deficiency is mostly caused by pertactin gene disruption by IS481. To assess the effect of the transition from whole-cell vaccine to ACV on the emergence of B. pertussis not expressing pertactin in Spain, we studied 342 isolates collected during 1986–2018. We identified 93 pertactin-deficient isolates. All were detected after introduction of ACV and represented 38% of isolates collected during the ACV period; 58.1% belonged to a genetic cluster of isolates carrying the unusual prn::del(–292, 1340) mutation. Pertactin inactivation by IS481 insertion was identified in 23.7% of pertactin-deficient isolates, arising independently multiple times and in different phylogenetic branches. Our findings support the emergence and dissemination of a cluster of B. pertussis with an infrequent mechanism of pertactin disruption in Spain, probably resulting from introduction of ACV.

Zanubrutinib for the Treatment of B-cell Malignancies

Selective Bruton's tyrosine kinase (BTK) inhibition is an important therapeutic approach for B-cell malignancies. Ibrutinib, a first-in-class, oral, irreversible BTK inhibitor, has a toxicity profile attributed to off-target inhibition of kinases structurally related to BTK. A highly potent and selective next-generation BTK inhibitor, zanubrutinib, was designed to address intolerance and toxicity concerns associated with ibrutinib. Zanubrutinib provides complete and sustained BTK occupancy in peripheral blood mononuclear cells and lymph nodes, with reduced toxicity compared with ibrutinib. Zanubrutinib received accelerated approval in the USA for patients with previously treated mantle cell lymphoma (2019) and relapsed/refractory (R/R) marginal zone lymphoma (2021) and was approved for Waldenström's macroglobulinaemia (WM) in 2021. Key clinical trials evaluating the efficacy and safety of zanubrutinib include the ASPEN study in patients with WM (ClinicalTrials.gov identifier: NCT03053440), the ALPINE study in patients with R/R chronic lymphocytic leukaemia (CLL) or small lymphocytic lymphoma (SLL; ClinicalTrials.gov identifier: NCT03734016) and the SEQUOIA study in treatment-naïve patients with CLL/SLL with and without del(17p) mutation (ClinicalTrials.gov identifier: NCT03336333). The more selective BTK binding profile, high potency, favourable pharmacokinetic and pharmacodynamic profile, and minimal cardiovascular toxicity of zanubrutinib compared with ibrutinib suggest that zanubrutinib may be an important treatment option when prescribed in accordance with guidelines.

Immunotherapy plus chemotherapy showed superior clinical benefit to chemotherapy alone in advanced NSCLC patients after progression on osimertinib.

BackgroundOsimertinib is the standard first‐line treatment for non‐small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) mutation. Resistance to osimertinib remains a clinical challenge. However, the optimal therapy for these patients is still controversial. In this study, we aimed to assess the efficacy and safety of immunotherapy plus chemotherapy (IO+C) compared with chemotherapy (C) in NSCLC patients after progression on osimertinib.MethodsAdvanced NSCLC patients after progression on osimertinib were retrospectively reviewed. Progression‐free survival (PFS), overall survival (OS), objective response rate (ORR), disease control rate (DCR), and safety were evaluated between the patients treated with IO+C and C.ResultsA total of 40 patients were included in the study. There were 20 patients each in the IO+C group or C group. The ORR was significantly higher in patients in the IO+C group (45% vs. 25%, p < 0.01). The median PFS was 6.4 months for patients in the IO+C group compared to 2.8 months for patients in C group (HR: 0.41, 95% confidence interval [CI]: 0.20–0.82, p < 0.01). The median OS was significantly longer in the IO+C group than the C group (OS: 12.8 vs. 10.5 months, HR: 0.39, 95% CI: 0.19–0.80, p < 0.01). In subgroup analysis, patients of both sexes, age ≤ 65, bone or adrenal metastasis, exon19 del mutation, and third‐line treatment obtained more OS benefits from immunotherapy. The safety profile of both groups was comparable.ConclusionsOur study provides the clinical evidence of favoring immunotherapy plus chemotherapy in NSCLC patients after progression on osimertinib.

Abstract 12102: TNNI3-K183del Mutation in Dilated Cardiomyopathy Patient-Specific iPSC-Derived Cardiomyocytes Affects Structural Proteins and Ion Channels, Leading to Impair the Function of Cardiomyocytes

Background: One of the causes for Dilated cardiomyopathy (DCM) are thought to be genetic defect. One mutation, Lys183 del mutation of the TNNI3, is suggested to be associated with left ventricular systolic dysfunction and transition from hypertrophic cardiomyopathy to DCM. However, the precise disease mechanism is not fully understood. Because cardiomyocytes derived from induced pluripotent stem cells (hiPSC-CMs) possess many of the same characteristics as primary human CMs and can be generated in unlimited quantities from pluripotent cell sources, they are thought to be a promising research tools to model genetic cardiomyopathies. Therefore, we hypothesized that iPS-CMs with mutations in TNNI3 is useful for elucidating the pathological mechanism of DCM and examined the phenotype of iPS-CMs with mutations in TNNI3. Methods and Results: We generated iPSCs from the patient with Lys183 del mutation of the TNNI3, and simultaneously induced iPS-CMs. By immunofluorescence stain analysis, we identified well-formed parallel arrays of repeating sarcomeres in myofibrils from WT iPS-CMs. On the other hand, iPS-CMs with Lys183 del mutation of the TNNI3 had fewer myofibrils and abnormal, irregular sarcomeres. In addition, sarcomere length was shorter in iPS-CMs with mutation in TNNI3 than in WT iPS-CMs. By gene expression analysis, we identified that iPS-CMs with mutation in TNNI3 had lower expression of some molecules including structural proteins or ion channels than WT iPS-CM. Cell motion analysis revealed that WT iPS-CMs were synchronously beating in multi wells, whereas iPS-CMs with mutation in TNNI3 were not. In addition, contraction velocity (CV) and relaxation velocity (RV) were significantly lower in iPS-CMs with mutation in TNNI3 (CV; 4.98±0.05 μm/sec, RV; 4.01±0.01 μm/sec) compared with WT iPS-CM (CV; 14.25±0.03 μm/sec, RV; 7.04±0.03 μm/sec). Moreover, significant decrease in contraction deformation distance and relaxation deformation distance were observed in iPS-CMs with mutation in TNNI3 (0.96±0.004 μm, 0.91±0.01 μm, respectively), compared to WT iPS-CMs (2.56±0.004 μm, 2.18±0.008 μm, respectively). Conclusion: iPS-CMs with Lys183 del mutation of the TNNI3 showed the morphological and functional phenotypes of DCM at a cellular level.

Real-World Persistence and Time to Next Treatment with Ibrutinib Treatment in Patients with Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma Including Patients at High Risk for Atrial Fibrillation or Stroke

Real-World Persistence and Time to Next Treatment with Ibrutinib Treatment in Patients with Chronic Lymphocytic Leukemia/Small Lymphocytic Lymphoma Including Patients at High Risk for Atrial Fibrillation or Stroke

Three-Year Follow-up of the Ascend Trial: Acalabrutinib Vs Rituximab Plus Idelalisib or Bendamustine in Relapsed/Refractory Chronic Lymphocytic Leukemia

Three-Year Follow-up of the Ascend Trial: Acalabrutinib Vs Rituximab Plus Idelalisib or Bendamustine in Relapsed/Refractory Chronic Lymphocytic Leukemia

SCREENING AND MOLECULAR CHARACTERIZATION OF HAEMOGLOBINOPATHIES USING ARMS (AMPLIFICATION REFRACTORY MUTATION SYSTEM) AND DIRECT DNA SEQUENCING TECHNIQUES AMONG YOUNG IN CENTRAL GUJARAT WESTERN INDIA

Haemoglobinopathies is consider the most common inherited disorders in human and results from genetic mutation in . one or more genes The present study was aimed to characterize the β-thalassemia mutation and haemoglobin variant in youth by ARMS PCR which is an uncomplicated and convenient method for identification of five common mutation from central Gujarat, western India region. This Study included 44 randomly selected haemoglobinopathies carrier student's sample of Anand People's Medicare Society (APMS), Anand for DNA analysis by ARMS PCR from March 2021-April 2021. Identification of five common Indian β thalassemia mutations along with Hb S and HB E were carried out by ARMS PCR method and δβ- thalassemia mutation was characterized by GAP-PCR. The samples which remain uncharacterized were sent to S N gene lab, Surat for DNA sequencing. In our study the most common mutation among five common mutations characterized was IVS-1, nt5 (G→C) in 22 (50%), followed by Codon41/42 (-CTTT) in 5 (11.3%). IVS-1, nt1 (G→T), Codon8/9 (+G) and 619bp del mutation was not identified in any carrier students screened for haemoglobinopathies. Other than these five common mutation Codon -88 (C→T) (2.27%) and Codon 30 (G→A) (2.27%) are also detected. The prevalence of haemoglobinopathies with respect to communities, reflects that SC/ST/OBC are at the highest risk with 50%. Communities like Rajput (22.7%), Patel (18.1%), Brahmin (6.8%) and Muslim (2.2%) are also showing prevalence. The study has included mutation in different communities reflects characterization of mutation of central Gujarat, western India which is significant for rapid and convenient identification of mutation while conducting screening programs and prenatal diagnosis. Rare mutations which are not recognized, need further confirmation for carrier detection followed by prenatal diagnosis.

Germline mutations in MSH2 and ATM gene in patients with GIST (gastrointestinal stromal tumor) and second epitelial tumors.

e23520 Background: In adult GISTs are frequently sporadic, while rarely GISTs are linked to Carney Triad and Carney-Stratakis Syndrome and NF1. GISTs with second primary tumors are reported in 4-33% of patients in literature and genetic counseling is suggested to explore an underlying germline mutations pathway. Methods: In our Academic Hospital Centre (EURACAN member) in Florence, Italy, we are following patients with GIST and multiple primary tumors with genetic counseling (72 GISTs with second tumors/185 patients with GIST) and germline analysis of the following genetic panel is performed as clinically indicated: BRCA1, BRCA2, MUTYH, MLH1, MSH2, MSH6, CDH1, ATM, TP53, PTEN, CHECK2, PALB2, BARD1, BRIP1, BLM, RAD51C, RAD51D, XRCC2, PMS2, MRE11A, RAD50, NBN, FAM175A, EPKAM, TSK1, MEN1 by sequencing analysis with Illumina MiSeq by kit multiplicom BRCA Hereditary cancer Mastr plus, and bioinformatic analysis by software SOPHIADDM (Sophia genetics) for point genetic alterations of BRCA1 NM_007294.3, BRCA2 NM_000059.3, MUTYH NM_000249, MSH2 NM_000251, MSH6 NM_000179, CDH1 NM_00444360, ATM NM_000051, TP53 NM_000546, PTEN NM_000314, CHEK2 NM_001005735, PALB2 NM_024675, BARD1 NM_000465, BRIP1 NM_032043, BLM NM_000057, RAD51C NM_002876, RAD51D NM_001142571, XRCC2 NM_005431, PMS2 NM_000535, MRE11A NM_005590, RAD50 NM_006732, NBN NM_002485, FAM175A NM_139076, EPCAM NM_002354, STK1 NM_000455, MEN1 NM_000244 and MLPA (Multiplex Ligation-dependent Probe Amplification) test analysis for patients with kit P087-BRCA1,P045-BRCA2(CHEK2, P248-MLH1-MSH2, P003-MLH1/MSH2, P072-MSH6-MUTYH (MRC-Holland). Results: In 3 patients germline mutations have been observed: 1 patient showed the c.1192dupG, p.(Ala398Glyfs*19) pathogenic mutation in exon 7 of MSH2 gene, confirmed by Sanger Sequencing, 1 patient showed c.565-?_1130+?del mutation consisting in heterozygous 3-4-5-6 exons deletion of MSH2 gene, confirmed by MLPA analysis, and in 1 patient the following ATM alteration has been identified in heterozygosis: ATM c.5319+2T &gt; C, p.(?). In the 2 patients with Lynch syndrome with colon adenocarcinoma (MSI-H), synchronous GISTs (1 patient quadruple WT and 1 patient kit ex 11 mutated ) were diagnosed; in the patient with ATM mutation, the diagnosis of GIST (kit ex 11 mutated) occurred after prostate adenocarcinoma and before colon adenocarcinoma (MSI-H). Conclusions: Our analysis suggests that GIST diagnosis could be tumor-related to multiple hereditary tumor syndromes as Lynch Syndrome and Ataxia-Teleangectasia syndrome, the latter being linked in eterozygosis to tumor susceptibility to breast in female. This report represents a high value in terms of genetic counseling for relatives and in terms of therapeutic implications for the patients.

Genomic variants in Fas-mediated apoptosis pathway predict a poor response to Platinum-based Chemotherapy for Chinese Gastric Cancer Patients.

Platinum-based adjuvant chemotherapy is very common for gastric cancer (GC) patients, but the chemotherapy sensitivity is very heterogeneous. The genomic variants and the gene-gene interactions involved in Fas-mediated apoptosis pathway including Fas (FAS 1377 G > A and 670 A > G), FasL (FASL 844 C > T) and caspase-8 (CASP8 -652 6N ins > del or I > D), may paly vital roles in the response to platinum-based treatment. In our investigation, 662 stage II-III postoperative GC patients were enrolled between 1998 and 2006. 261 patients accepted platinum-based regimens and the remaining 401 were not. The log rank tests, Kaplan Meier plots, Pearson chi-square tests, Student t-tests and Cox regression analyses were performed. For the chemotherapy cohort, FAS 1377 G > A or FAS 670 A > G variants alone was related with inferior survival, and a greater than additive effect was identified when patients simultaneously carrying FAS 1377 GA and FAS 670 GA genotypes. But the poor response was neutralized when patients simultaneously carrying FASL 844 C > T or CASP8 -652 6N ins > del mutations. Our study suggested that FAS 1377 G > A and FAS 670 A > G variants may serve as potential biomarkers to predict the response to platinum-based adjuvant chemotherapy, and the gene-gene interactions involved in Fas-mediated apoptosis pathway may enhance or neutralize the chemosensitivity.