Del Mutation Research Articles

Detection of epidermal growth factor receptor mutation in the peripheral blood of patients with esophageal carcinoma to guide epidermal growth factor receptor-tyrosine kinase inhibitor treatment.

This study aimed to explore the epidermal growth factor receptor (EGFR) mutation in the peripheral blood of patients with esophageal carcinoma and analyze the relationship between EGFR-tyrosine kinase inhibitor (EGFR-TKI) therapeutic effect and EGFR mutation in peripheral blood. A retrospective analysis was performed on 66 patients with esophageal carcinoma treated with EGFR-TKI (Gefitinib) from February 2014 to March 2017 in our hospital. Real-time polymerase chain reaction was applied to detect the mutation of the EGFR gene in peripheral blood specimens before patients were treated with EGFR-TKI. The relationship between EGFR mutation in the peripheral blood and clinical features of patients were analyzed. The correlation between EGFR mutation in peripheral blood and EGFR-TKI treatment response was also demonstrated. Among the 66 patients, 18 cases were determined as EGFR mutation in peripheral blood. The mutation rate was 27.3%. Among these patients, it observed 1 case of exon 18 2155G>A mutation, 3 cases of exon 19 2235-2249Del mutation, 5 cases of 2236-2250Del mutation, 1 case of 2254-2277Del mutation, 1 case of L747-A750Del mutation, 3 cases of exon 21 2576T>G mutation, 3 cases of 2497T>G mutation, and 1 case of 2504A>T mutation. EGFR mutation in the peripheral blood of patients with esophageal carcinoma was unrelated to the gender, age, and the location of the tumor (P < 0.05). By contrast, EGFR mutation was related to the pathological types of the patients (P < 0.05). The mutation rate of the EGFR of squamous cell carcinoma patients was significantly higher than that of adenocarcinoma patients (P < 0.05). Among EGFR wild-type patients, 8 cases were stable disease (SD), and 40 cases were progressive disease (PD), after EGFR-TIK treatment. No complete response (CR) and partial response (PR) patients were reported. The overall response rate (ORR) was 0.0%. Among the EGFR mutation group, 5 were SD cases, 5 were PD cases, and 8 were PR cases. No CR case was reported. The ORR was 44.4%. The ORR of the EGFR group was significantly higher than that of the wild-type group (P < 0.05). The detection of EGFR mutation in peripheral blood can be applied as an effective index for EGFR-TKI (Gefitinib) treatment in patients with esophageal carcinoma.

BENDAMUSTINE: AN OLD DRUG IN THE NEW ERA FOR PATIENTS WITH NON-HODGKIN LYMPHOMAS AND CHRONIC LYMPHOCYTIC LEUKEMIA.

SUMMARY – The aim of this review is to present data on bendamustine, a non-cross resistant alkylating agent, alone or in combination for treatment of non-Hodgkin lymphoma (NHL) and chronic lymphocytic leukemia (CLL). Bendamustine is currently approved for rituximab-resistant indolent NHL and CLL in patients not fit for conventional chemotherapy. Recent studies have shown superiority of bendamustine combination with rituximab (B-R) in first line treatment of indolent NHLs and mantle cell lymphoma, suggesting a shift of the standard of care in this setting. B-R regimen has also shown efficacy in relapsed setting suggesting the possible treatment option for patients failing conventional chemotherapy. In rituximab-resistant NHL, the recent GADOLIN study exploring the addition of obinutuzumab to bendamustine has yielded impressive result changing the standard of care in this hard-to-treat population. Concerning CLL, despite inferiority to the standard of care in young fit patients, as defined in CLL10 study, B-R has yielded a more beneficial toxicity profile and its use in first line treatment should be decided individually. In relapsed setting, the addition of ibrutinib to B-R has shown superior results compared to B-R alone, possibly changing the paradigm of treatment of relapsed CLL. In conclusion, bendamustine as a single agent or in combinations has shown activity with acceptable toxic profile in the treatment of patients with indolent NHLs or CLL without del(17p) mutation.

Investigation of the Connexin-30 gene Deletion Mutations Del (GJB6-D13s1830) and Del (GJB6-D13s1854) among Subjects with Non-Syndromic Sensorineural Hearing Loss in Khorasan Razavi Province

Introduction: Hearing loss (HL) is the most common sensorineural disorder with the frequency of 1 to 1000 neonates and it's seen in two syndromic and non-syndromic states. Given that the mutations of DFNB1 locus, containing Connexin 26 (Cx26) and 30 (Cx30) genes are the only responsible for 50% non-syndromic HL with autosomal recessive hereditary in different populations, the aim of this study is the screening of the connexin30 gene deletion mutations including Del (GJB6-D13S1830) and Del (GJB6-D13S1854) in HL subjects were negative to mutation in Cx26 gene. Materials & methods: In this descriptive study, 85 unrelated patients (44 males and 41 females) with autosomal recessive hearing loss of Khorasan Razavi province that were negative towards the Cx26 gene mutations, were studied. Genomic DNA was extracted from 1 ml of EDTA anticoagulated peripheral blood by rapid genomic DNA extraction (RGDE) method. For molecular analysis, multiplex PCR and direct sequencing methods were used. Findings: Out of 170 studied chromosomes, none of the Del (GJB6-D13S1830) and Del (GJB6-D13S1854) mutations found in the studied subjects. Discussion & conclusions: Lack of the deletion mutations in the Cx30 gene, indicates the involvement of other genes in causing disease. Thus, it is necessary to study other HL genes in these subjects.

Screening for DEL phenotype in RhD negative Indians.

DEL phenotype represents a very weak form of D variant detected only by adsorption and elution technique. DEL phenotype individuals mistyped as RhD-negative can lead to alloimmunization after transfusion or pregnancy. Molecular techniques have now been used to identify DEL variants. They are commonly encountered in the East Asian population with RHD(K409K) being the most frequent allele. RHD(M295I) is the most common DEL allele in Caucasians. As there is a paucity of data on DEL phenotype in the Indian population, the study aims to screen RhD negative individuals for two most common DEL mutations. EDTA blood was collected from 900 RhD negative individuals. Serological analysis included testing for the five major Rh antigens- C, c, D, E, and e by tube technique. Samples showing negative reaction for the presence of D antigen by Indirect Antiglobulin test were further tested for DEL phenotype by adsorption and elution technique. Molecular analysis involved DNA extraction and testing by PCR-SSP for RHD(K409K) and RHD(M295I) DEL alleles. Rh phenotyping showed 153 Rh negative individuals with r'r, ten with r''r and 737 with rr phenotype. All the samples tested negative for RhD antigen by adsorption and elution method. The two common DEL mutations RHD(K409K) and RHD(M295I) were also not detected in the study population. The study population showed the absence of the two common DEL alleles, concluding the variant to be rare. A comprehensive study with a larger sample size to look for other DEL mutations should be performed.

Differential Analysis of Genetic, Epigenetic, and Cytogenetic Abnormalities in AML.

Acute myeloid leukemia (AML) is a haematological malignancy characterized by the excessive proliferation of immature myeloid cells coupled with impaired differentiation. Many AML cases have been reported without any known cytogenetic abnormalities and carry no mutation in known AML-associated driver genes. In this study, 200 AML cases were selected from a publicly available cohort and differentially analyzed for genetic, epigenetic, and cytogenetic abnormalities. Three genes (FLT3, DNMT3A, and NPMc) are found to be predominantly mutated. We identified several aberrations to be associated with genome-wide methylation changes. These include Del (5q), T (15; 17), and NPMc mutations. Four aberrations—Del (5q), T (15; 17), T (9; 22), and T (9; 11)—are significantly associated with patient survival. Del (5q)-positive patients have an average survival of less than 1 year, whereas T (15; 17)-positive patients have a significantly better prognosis. Combining the methylation and mutation data reveals three distinct patient groups and four clusters of genes. We speculate that combined signatures have the better potential to be used for subclassification of AML, complementing cytogenetic signatures. A larger sample cohort and further investigation of the effects observed in this study are required to enable the clinical application of our patient classification aided by DNA methylation.

P3.02b-041 Genome-Wide Screen of DNA Methylation Changes Reveals GABBR2 as a Novel Potential Target for EGFR 19 Deletion Adenocarcinoma with Erlotinib: Topic: EGFR Biomarkers

Lung cancer is the leading cause of cancer related death around the world. The last decade has witnessed the rapid development of epidermal growth factor receptor (EGFR) directly targeted therapies that have significantly changed the treatment of non-small-cell lung cancer (NSCLC). However, the challenge of acquired resistance to EGFR tyrosine kinase inhibitors (TKIs) has been an issue when talking with activating EGFR mutations, which makes it crucial to elucidate the mechanism of EGFR-TKI targeted drug resistance. Methyl-sensitive cut counting sequencing (MSCC), one of the most commonly used whole-genome DNA methylation sequencing technology, was applied to investigate the changes of paired tissue DNA methylation before and after TKI (erlotinib) treatment lasting two cycles with partial response (PR) for stage IIIa (N2) lung adenocarcinoma patients (N=2) with activating EGFR 19 deletion. Sequenom EpiTYPER assay method was further analyzed to double confirm the changed methylated candidate genes in these two patients, through comparing the methylated changes in paired tissues before and after TKI treatment. Western blotting, cell cycle and apoptosis analysis by the up/down regulation of a candidate gene (GABBR2) were performed in three lung adenocarcinoma cell lines, A549 (EGFR wild type), HCC4006 (EGFR 19 deletion, DelL747-E749) and HCC827 (EGFR 19 deletion, DelE746-A750), to elucidate the mechanism of GABBR2 gene in the regulation of EGFR-TKI treatment. Sixty aberrant methylated genes were firstly discovered using MSCC method in these two patients harboring EGFR 19 del mutation treated with erlotinib. Two aberrant methylated genes, CBFA2T3 and GABBR2, were clearly validated by the Sequenom EpiTYPER assay subsequently. GABBR2 was significantly down regulated in HCC4006 and HCC827 cells harboring EGFR 19 mutations but remained conservative in A549 cells with EGFR wild-type after erlotinib treatment by Western blotting. The phenomenon was in line with the obvious apoptosis of HCC4006 and HCC827 cell lines after erlotinib treatment, but not in A549 cells. GABBR2 was further induced down regulation after erlotinib exposure through apoptosis method silenced by siRNA using RNAi technology. Meanwhile, GABBR2 gene was demonstrated up regulation rescued the apoptosis significantly, when overexpressing GABBR2 in HCC827 cell lines along with erlotinib treatment. Genome-wide screen of DNA methylation changes demonstrated that GABBR2 gene might be as a novel potential treatment target for stage IIIa (N2) EGFR 19 deletion adenocarcinoma with erlotinib treatment. Our research provides a new theoretical basis for the epigenetic study of EGFR mutated lung adenocarcinoma treatment.

Effect of Lenalidomide (LEN) Exposure on Response and Outcomes in Patients (Pts) with Lower-Risk Non-Del(5q) Myelodysplastic Syndromes (MDS)

▪Introduction: In the phase 3 MDS-005 study, LEN treatment was associated with a significant increase in transfusion independence (TI) vs placebo (26.9% vs 2.5%; P < 0.001) (Santini et al. J Clin Oncol. 2016;pii:JCO.2015.66.0118) and greater clinical benefit vs placebo (31.9% vs 3.8%) (Garcia-Manero et al. Haematologica. 2016;101:abstract P252) in red blood cell (RBC) transfusion-dependent (TD) pts with lower-risk non-del(5q) MDS who were ineligible for or refractory to erythropoiesis-stimulating agents (ESAs). For pts with lower-risk del(5q) MDS, a starting dose of LEN 10 mg is recommended, followed by subsequent dose modifications in response to adverse events (AEs), in order to maximize duration of treatment and thereby improve pt outcomes. This study evaluated the relationship between LEN exposure, including dose reductions, and duration of treatment, and the clinical benefit to pts with lower-risk, non-del(5q) MDS in the multicenter, randomized, double-blind, phase 3 MDS-005 study.Methods: Eligible pts were age ≥ 18 years, had TD anemia due to International Prognostic Scoring System (IPSS)-defined Low- or Intermediate-1-risk MDS, lacked the del(5q) mutation, and were ineligible for or refractory to ESAs. Pts were randomized 2:1 to receive LEN 10 mg (n = 160) or placebo (n = 79) once daily (both in 28-day cycles). Pts randomized to LEN with creatinine clearance ≥ 40 to < 60 mL/min (n = 57) received LEN 5 mg. Cumulative dose, dose reductions, and discontinuations were recorded; association between these parameters and clinical benefit or RBC-TI was evaluated. Clinical benefit ≥ 8 weeks was defined as the composite endpoint of RBC-TI ≥ 8 weeks, or transfusion reduction of ≥ 4 units packed RBCs (pRBCs) ≥ 8 weeks, or hemoglobin (Hb) increase ≥ 1.5 g/dL ≥ 8 weeks (International Working Group 2006), or cytogenetic response (CyR). Rate of transfusion reduction was calculated using data collected during a 112-day assessment period to account for on-study differences in transfusion burden. LEN dose modifications due to AEs were defined in the study protocol. The analysis included all pts receiving LEN.Results: Of 160 LEN-treated pts included in the analysis, 77 (48.1%) underwent ≥ 1 LEN dose reduction. A total of 73 pts experienced ≥ 1 dose reduction due to AEs, amounting to 102 dose reductions overall. Of these AE-related dose reductions, 47 were due to neutropenia, 33 due to thrombocytopenia, and 2 due to other hematologic conditions. Median time to first dose reduction was 85 days (interquartile range [IQR] 50-114 days). A total of 145 LEN-treated pts discontinued therapy: 52 discontinued due to AEs; 32 discontinued before the end of cycle 3, of which 23 were due to AEs.Of the 77 pts who had ≥ 1 dose reduction, 6 (7.8%) pts had a dose reduction in the first cycle; 37 (48.1%) pts had dose reductions during cycles 1-3. During the first 3 cycles of LEN, pts with ≥ 1 dose reduction received a lower median dose than those with no dose reductions (385 mg [IQR 318-520 mg] vs 660 mg [IQR 415-840 mg]). However, when analyzed over the course of the entire study, pts undergoing ≥ 1 dose reduction received a higher median cumulative dose than those with no dose reductions (950 mg [IQR 595-1,435 mg] vs 715 mg [IQR 370-1,660 mg]). Duration of treatment was also longer for pts with ≥ 1 dose reduction vs those with no dose reductions (172 days [IQR 140-391 days] vs 92 days [IQR 46-168 days]).Pts with ≥ 1 dose reduction were more likely to achieve RBC-TI ≥ 8 weeks than pts with no dose reductions (39% vs 16%; odds ratio [OR] 3.44 [95% confidence interval {CI} 1.63-7.26]). Pts with ≥ 1 dose reduction were also more likely to achieve clinical benefit ≥ 8 weeks vs pts with no dose reductions (47% vs 18%; OR 3.98 [95% CI 1.94-8.15]).Conclusions: In this post hoc analysis of the MDS-005 study, pts undergoing ≥ 1 LEN dose reduction had a longer duration of treatment, and were more likely to achieve RBC-TI and the composite endpoint of clinical benefit (RBC-TI ≥ 8 weeks, transfusion reduction ≥ 4 units pRBCs ≥ 8 weeks, Hb increase ≥ 1.5 g/dL ≥ 8 weeks, or CyR) than pts with no dose reductions. DisclosuresSantini:Novartis: Consultancy, Honoraria; Onconova: Other: advisory board; Amgen: Other: advisory board; Janssen: Consultancy, Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Astex: Other: advisory board. Almeida:BMS: Speakers Bureau; Novartis: Consultancy, Speakers Bureau; Celgene: Consultancy, Research Funding, Speakers Bureau; Alexion: Speakers Bureau; Shire: Speakers Bureau. Fenaux:Celgene, Novartis, Teva: Honoraria; Celgene, Janssen, Novartis, Astex, Teva: Research Funding. Gatterman:Celgene Corporation: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Heinrich-Heine-Universitat Dusseldorf: Employment. Ozawa:Sumitomo Dainippon Pharma Co. Ltd: Research Funding; JCR Pharmaceutical Inc: Consultancy; Celgene Japan: Consultancy; Takara Bio Inc: Research Funding. Goldberg:COTA Inc: Employment; Novartis: Consultancy; Pfizer: Honoraria; Bristol Myers Squibb, Novartis: Speakers Bureau; Neostem: Equity Ownership. Weaver:Celgene Corporation: Employment, Equity Ownership. Sugrue:Celgene Corporation: Employment, Equity Ownership.

First report of prevalence c.IVS1+1G>A and del (GJB6-13S1854) mutations in Syrian families with non-syndromic sensorineural hearing loss

ObjectiveMutations in GJB2 and GJB6 genes are a frequent cause of congenital non-syndromic hearing loss (NSHL). Mutational screening has usually focused on coding region of GJB2 gene. A few studies have been conducted on the non-coding region and exon 1. c.IVS1+1G>A (a splice site mutation in GJB2 gene have been detected as disruptive mutation. Del (GJB6 D13S1830) is found in many populations, but del (GJB6 D13S1854) is reported from a few restricted countries. This study was carried out to investigate the prevalence of splice site mutation c.IVS1+1G>A and two common deletions in GJB6 gene as the genetic etiology of hearing impairment in 70 Syrian families. MethodsThe frequency of the c.IVS1+1G>A mutation and two deletions were determined by PCR-RFLP and A multiplex PCR assay. ResultOur results showed a high prevalence of IVS1+1G>A mutation (20%) and del(GJB6-D13S1854) (15.7%) in deaf families. The homozygous genotype (c.IVS1+1G>A/c.IVS1+1G>A) was observed in one family and the compound heterozygous genotypes (c.35delG/c.IVS1+1G>A) and (c.IVS1+1G>A/V153I) were observed in 7 families and one family respectively. Also, the heterozygous state (c.IVS1+1G>A/unknown) was detected in 5 families. The study of del((GJB6-D13S1854) was showed a compound heterozygous genotype del((GJB6-D13S1854)/c.IVS1+1G>A) in the same families (5 families) having heterozygous genotype of c.IVS1+1G>A mutation. Also, del(GJB6-D13S1854) is combined with c.35delG mutation in 2 families and it was observed in the heterozygous state del(GJB6-D13S1854)/unknown) in 4 families. In contrast, the del(GJB6-D13S1830) described in many population was absent in our patients. ConclusionOur findings indicate to significant contribution of the splice site mutation and del(GJB6-D13S1854) in our deaf families and these mutations were important causes of hearing impairment.

Severe left ventricular hypertrophy and marked cardiac fibrosis in Danon disease

A seventeen year-old male underwent orthotopic cardiac transplantation after development of heart failure with progression to New York Heart Association (NYHA) Class III symptoms over six months. He had a known diagnosis of Danon disease confirmed on previous genetic testing, identified as the LAMP-2 Xq24 G14 del mutation. There was an extensive family history, with his maternal grandmother, mother, two uncles and older brother all previously diagnosed with this condition. His two uncles had died of sudden cardiac death (SCD) as young adults. His eighteen year-old brother died of worsening cardiac failure and refractory ventricular fibrillation one year previously while awaiting transplantation. His mother also had a dilated cardiomyopathy, with a secondary prevention internal cardioverter defibrillator (ICD) in situ for survived ventricular fibrillation.

Bevacizumab to combat EGFR-TKI resistance in a patient with advanced non-small cell lung cancer harboring an EGFR mutation: A case report.

Treatment with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) is the first-line strategy for patients with non-small cell lung cancer (NSCLC) harboring EGFR-activating mutations. Acquired resistance to EGFR-TKIs is inevitable in patients receiving EGFR-TKI therapy. Treatment with bevacizumab can induce a marked improvement in the overall and progression-free survival of patients with NSCLC; however, the effect of bevacizumab on TKI resistance in patients with NSCLC with an activating EGFR mutation is largely unknown. The present study reports the case of a patient with advanced, metastatic lung adenocarcinoma harboring 19 Del mutations, and who developed resistance to afatinib. The addition of bevacizumab to afatinib treatment was shown to overcome the acquired TKI resistance in the patient, as well as to promote an improved outcome for her brain metastases.

Clinical features and gene mutation analysis of cartilage oligomeric matrix protein in a family with pseudoachondroplasia

Objective To investigate cartilage oligomeric matrix protein(COMP)gene mutation in a three-generation pedigree with two cases of pseudoachondroplasia, and to definitize genotype-phenotype correlation. Methods The clinical data and peripheral blood were collected from the patients with pseudoachondroplasia and their family members. All the 19 exons and their flanking sequences of COMP gene in two patients and three unaffected family numbers and 50 unrelated individuals were analyzed by PCR amplification and direct sequencing. Results The proband, a 6-year-old girl presented with typical clinical features of pseudoachondroplasia, including disproportionate short limb dwarfism, staggering gait, double genu varus deformity, and wider clinical and imaging long bone metaphysis. The 33-year-old father showed a similar manifestation including disproportionate short limb dwarfism and double genu varus deformity, and was performed correcting operation on lower limbs for double genu varus at the age of 10 years. DNA sequencing analysis of the COMP gene revealed a del mutation(c.1417_1419delGAC)in exon 13 in two patients with pseudoachondroplasia, but not in the other unaffected members from the pedrgree and 50 control subjects. Conclusion A del mutation c. 1417_1419delGAC of COMP gene may contribute to the disease in the pedigree. Key words: Pseudoachondroplasia; Cartilage oligomeric matrix protein; Gene mutation

Two cases of familial cerebral cavernous malformation caused by mutations in the CCM1 gene.

Cerebral cavernous malformation (CCM) is a vascular malformation characterized by abnormally enlarged capillary cavities without any intervening neural tissue. We report 2 cases of familial CCMs diagnosed with the CCM1 mutation by using a genetic assay. A 5-year-old boy presented with headache, vomiting, and seizure-like movements. Brain magnetic resonance imaging (MRI) revealed multiple CCM lesions in the cerebral hemispheres. Subsequent mutation analysis of his father and other family members revealed c.940_943 del (p.Val314 Asn315delinsThrfsX3) mutations of the CCM1 gene. A 10-month-old boy who presented with seizure-like movements was reported to have had no perinatal event. His aunt was diagnosed with cerebral angioma. Brain and spine MRI revealed multiple angiomas in the cerebral hemisphere and thoracic spinal cord. Mutation analysis of his father was normal, although that of the patient and his mother revealed c.535C>T (p.Arg179X) mutations of the CCM1 gene. Based on these studies, we suggest that when a child with a familial history of CCMs exhibits neurological symptoms, the physician should suspect familial CCMs and consider brain imaging or a genetic assay.

Investigation of the GJB6 Deletion Mutations Del (GJB6-D13s1830) and Del (GJB6-D13s1854) in Iranian Patients with Autosomal-Recessive Non-Syndromic Hearing Loss (ARNSHL)

Hearing loss (HL) is the most common inherited sensory disorder affecting about 1 in 1000 births. The first locus for nonsyndromic autosomal recessive HL is on chromosome 13q11-22. The two genes, GJB2 and GJB6, are closely located on chromosome and are known to be co-expressed in the embryonic cochlea. Deletion mutations involving GJB6 were associated with autosomal-recessive nonsyndromic hearing loss (ARNSHL) and in combination with a GJB2 mutation with digenic ARNSHL. The objective of this study was to screen for the del (GJB6-D13S1830) and del (GJB6-D13s1854) mutations in GJB6 gene in patients with ARNSHL from Iran, using multiplex PCR and direct sequencing methods. Agarose gel electrophoresis and DNA sequencing of amplified fragment of the PCR reaction showed none of the patients was found to carry deletion in GJB6 gene which indicates that these deletions are restricted to certain populations and indicating a founder effect regarding these deletions.

A Phase II Study of Ofatumumab-High Dose Methylprednisolone Followed By Ofatumumab-Alemtuzumab in 17p Deleted or TP53 Mutated CLL

Abstract BACKGROUND: Even in the age of kinase inhibitors, the prognosis for patients (pts) with CLL with del(17p) or TP53 mutations remains poor, with the median PFS only 28 mo. for pts with relapsed/refractory del(17p) CLL treated with ibrutinib. High dose methylprednisolone (HDMP) and alemtuzumab (alem) both have activity in 17p disease and work independently of TP53. Prior work has demonstrated that giving HDMP/alem simultaneously is efficacious, but toxic. We hypothesized that giving these agents sequentially with ofatumumab (ofa) would maintain efficacy while decreasing toxicity. METHODS: This phase II study of ofa/HDMP followed by ofa/alem in CLL pts with 17p deletion or TP53 mutation employed a parallel 2-arm design (treatment-naive (TN) and relapsed/refractory (R/R)). Therapy was the same in both arms and included ofa/HDMP for 2-4 cycles (part A) followed by ofa/alem for 4-24 weeks, up to maximum response (part B). Responders could proceed to alloHSCT or a maintenance phase with ofa given q2 mo and alem given q2 wks (part C). Antimicrobial prophylaxis for PCP, HSV/VZV, and fungal infections was mandatory, as was G-CSF support. The primary objective was to estimate the ORR at the conclusion of the two-part induction therapy in both cohorts. Secondary objectives were to estimate the rate of CR, objective response by compartment, rate of MRD negativity by 4 color flow cytometry, PFS, and OS, rate at which transplant-eligible pts were able to proceed to alloHSCT, and to assess safety. Toxicity was assessed by IW-CLL and CTCAE v4.0. Response assessments by IW-CLL criteria were performed mid-way and at the end of parts A and B, and q6 mo. on part C. RESULTS: A total of 30 patients were enrolled. Baseline pt characteristics were as follows: TN (n=15): median age 64 (range 45-86), median WBC 58K, Hct 31, Plts 134, B2M 4.3, IGHV unmutated 79%, median %bone marrow (BM) involvement 80%. R/R (n=15): median age 65 (range 58-80), median WBC 31, Hct 33, Plts 114, B2M 4.5, IGHV unmutated 73%, median %BM involvement 60%, median # prior therapies 2 (range 1-4) including 9 pts with prior FCR or FR and 7 pts with prior BR. One patient in each arm had TP53 mutation without 17p deletion, and 4 pts had mut NOTCH1. The median number of copy number changes by SNP array was high, at 12.4 in the TN and 14 in the R/R cohorts. In the TN arm, 14/15 pts moved from part A-&gt;B, 5 moved from part B-&gt;C, and 5 moved on to alloHSCT in remission. The best ORR for the TN arm was 80% (67% PR, 13% CR), with 12/15 (80%) pts achieving BM MRD negativity. The TN 2-yr PFS and OS are 70% and 85%, respectively. In the R/R arm, 8/15 pts moved from part A-&gt;B, 4 moved from part B-&gt;C, and 5 moved on to alloHSCT in remission. The best ORR for the R/R arm was 68% (all PRs), with 8/15 (54%) pts achieving BM MRD negativity. The R/R 2-yr PFS and OS are 53% and 67%, respectively. Responses in both arms were independent of TP53 and NOTCH1 mutation status. Studywide, a greater number of somatic mutations was associated with shorter PFS and OS (HR 1.13 per mutation, 95% CI 1.02-1.25, p=0.015 and HR 1.185, 95% CI 1.047-1.341, p=0.0073, respectively). The most common grade 3/4 toxicities were: neutropenia (33%), thrombocytopenia (20%), anemia (10%). Infectious complications included pneumonia (5 cases, 1 Gr2, 3 Gr3, 1 Gr4), febrile neutropenia (2 cases, both Gr3), and cellulitis (1 case, grade 3). Four pts had low-level CMV reactivation, but no CMV infections occurred. Venous thromboembolism occurred in 3 pts. With a median follow-up time among survivors of 25 mo., 21 of the total 30 pts are still alive, and causes of death included: progressive disease n=6, infection n=3. 12/18 (67%) pts who were transplant eligible were able to proceed to HSCT. Discontinuations were due to: progressive/refractory disease (n=10, including 3 Richter's transformations (R/R n=2, TN n=1)), physician decision (n=2), and unacceptable toxicity (n=1). CONCLUSION: Ofa/HDMP followed by Ofa/alem is highly active for both TN and R/R CLL pts with del(17p) and/or TP53 mutation. In addition to a robust ORR, we observed a high rate of MRD-negativity in the bone marrow, which allowed most of the transplant-eligible pts to proceed to alloHSCT. Sequential dosing appears to reduce infectious complications compared to concurrent dosing. This regimen is a feasible option for pts with ultra-high risk CLL to facilitate maximal cytoreduction prior to alloHSCT. &lt;&gt; Disclosures Davids: Genentech: Other: ad board; Pharmacyclics: Consultancy; Janssen: Consultancy.

Treatment Selection and Practice Patterns for the Management of High-Risk Chronic Lymphocytic Leukemia (CLL) in the US: An Analysis of the Impact of Risk Stratification on Treatment Selection from the Connect CLL® Registry

Treatment Selection and Practice Patterns for the Management of High-Risk Chronic Lymphocytic Leukemia (CLL) in the US: An Analysis of the Impact of Risk Stratification on Treatment Selection from the Connect CLL® Registry

Abstract C80: Development of EGFR C797S mutation in serial liquid biopsy assessments in the clinical practice setting

Abstract Introduction: EGFR T790M has emerged as an important biomarker to predict response to 3rd-generation tyrosine kinase inhibitors (TKI). Therapies such as AZD 9291 and Rociletinib demonstrated impressive results in the clinical setting in patients with non-small cell lung cancer (NSCLC). However just as this resistance biomarker (T790M) has developed as a result of treatment with prior EGFR TKIs, EGFR C797S mutation is an acquired resistance mechanism to EGFR T790M inhibitors. This mutation was recently reported for the first time in clinical practice utilizing tissue-biopsy based next generation sequencing (NGS) and in the clinical trial setting in cell-free circulating DNA (cfDNA). Here we report the first case of a patient with NSCLC and emergence of C797S mutation utilizing a cfDNA biopsy-free NGS panel in serial assessments in the clinical setting. Methods: Patients with NSCLC were seen at Levine Cancer Institute and had serial blood draws to assess molecular aberrations in cfDNA over time for comparison with radiographic reviews. Blood samples were sent to Guardant360 for assessment by NGS with a targeted cfDNA NGS panel for 68 genes with complete exon sequencing for all 28 exons in EGFR, and other genes in the panel. Mutant allele fractions (MAF) are reported as% of mutant DNA molecules divided by total molecules (mostly leukocyte DNA-derived) overlapping the same mutated nucleotide base position. Results: A 50 year old African-American female never-smoker with lung adenocarcinoma developed a tissue-biopsy confirmed EGFR T790M mutation after 12 months on erlotinib for EGFR exon 19 deletion. She had significant clinical and radiographic responses to a 3rd generation TKI started May 2014. cfDNA NGS tests in March and April 2015 showed no genomic alterations detectable. In late May 2015, she developed clinical symptoms of cough, dyspnea and fatigue but no definitive radiographic evidence of progression on scans June 2015. She received steroids and antibiotics and continued treatment with the 3rd generation TKI. However, corresponding cfDNA NGS testing revealed emergence of multiple low level mutations and the original deletion in EGFR (EGFR L747_P753 DelInsS, T790M, C797S, and A755G variant of uncertain significance) and TP53 (all &amp;lt;1.1%). Repeat scans in July 2015 demonstrated clear progression of disease and a fourth cfDNA NGS test showed a rise in all low level aberrations previously detected in cfDNA: EGFR A755G 4.2%, TP53 P278R 3.6%, EGFR T790M 3.1%, EGFR L747_P753 DelInsS 2.9%, EGFR C797S 1.4%. NGS indicated that there may be 2 clones present for EGFR C797S but confirmation is required as disease progresses. Discussion: Complete exon sequencing of cfDNA upon clinical progression uncovered the simultaneous re-emergence of the original driver EGFR exon 19 del and resistance driver mutation EGFR T790M as well as the emergence of the next resistance driver mutation EGFR C797S. The latter mutation interferes with covalent binding of a 3rd generation TKI and under treatment pressure, the tumor appears to have simultaneously evolved two different EGFR C797S clones at low MAFs. cfDNA can be a dynamic measure of tumor response, easily obtained from a patient on any visit with potential for detection before macroscopic/radiographic evidence of progression, as demonstrated in this case with tumor evolution of the EGFR T790M and C797S alterations. Citation Format: Kathryn F. Mileham, Qing Zhang, Carol J. Farhangfar, Daniel E. Haggstrom, Stephen Fairclough, Oliver A. Zill, Daniel R. Carrizosa, Richard B. Lanman, Edward S. Kim. Development of EGFR C797S mutation in serial liquid biopsy assessments in the clinical practice setting. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C80.

Prevalence of 35de1G/GJB2 and del (GJB6-D13S1830) mutations in patients with non-syndromic deafness from a population of Espírito Santo - Brazil

Mutations in GJB2 gene are the leading cause of deafness in autosomal recessive inheritance, and the 35delG mutation is the most common in many ethnic groups. Besides the 35delG mutation in homozygosis, the mutation is also found in compound heterozygosis, coupled with other mutations in genes GJB2 and GJB6. AimTo determine the prevalence of 35delG/GJB2 and del (GJB6-D13S1830) mutations in patients with sensorineural hearing impairment in residents from the Espirito Santo state, Brazil. Materials and methods77 unrelated individuals with moderate to profound sensorineural hearing loss were evaluated. The 35delG mutation was studied by PCR / RFLP; and the del (GJB6-D13S1830) mutation was screened by the technique of multiplex PCR. Results88.3% had normal genotype for the studied mutations, 1.3% were compound heterozygotes, 3.9% homozygotic for the 35delG mutation, 6.5% heterozygotic for 35delG/GJB2. The frequency of 35delG/GJB2 and del (D13S1830/GJB6) alleles in the sample was 7.8% and 0.65%, respectively. ConclusionThe data confirmed the existence of the mutations studied in cases of sensorineural hearing loss in a population from Espírito Santo / Brazil. These findings reinforce the importance of genetic diagnosis, which can provide early treatment for children and genetic counseling for the affected families.

A Novel Case of Myelodysplastic Syndrome With Isolated del(5q) and Concomitant CALR Mutation

Myelodysplastic syndromes (MDSs) and myeloproliferative neoplasms (MPNs) are subgroups of myeloid neoplasms with chronic clinical courses. MDSs are characterized by simultaneous proliferation and apoptosis of hematopoietic cells and peripheral blood cytopenias. MDS with isolated del(5q) is a specific subtype of MDS that occurs primarily in older women, and is characterized by macrocytic anemia, increased platelet count, micromegakaryocytes, and favorable clinical course. …

Screening of GJB6 gene large deletions among Syrians with congenital hearing impairment.

Research into the genetics of congenital hearing impairment in the Syrian population, where cases are noticeably encountered, is still in its infancy. Our goal was to estimate the frequencies of the del(GJB6-D13S1830) and del(GJB6-D13S1854) mutations in a group of Syrians with autosomal recessive nonsyndromic hearing loss (ARNSHL). Forty-one unrelated Syrian probands, already screened for exon 2, GJB2 gene mutations, were reanalyzed for del(GJB6-D13S1830) and del(GJB6-D13S1854) mutations by polymerase chain reaction. The del(GJB6-D13S1830) mutation was only found in homozygosity in 1 of 41 probands (2.43%), while the del(GJB6-D13S1854) mutation was not detected in any of the enrolled patients. Coexistence of GJB2 and GJB6 mutations was not encountered in any case. Our study reports the first case in Syria with the del(GJB6-D13S1830) mutation. This mutation might be considered in the diagnosis and genetic counseling of inherited hearing impairment in the Syrian population.

Bladder Cancer and Genetic Mutations.

The most common type of urinary bladder cancer is called as transitional cell carcinoma. The major risk factors for bladder cancer are environmental, tobacco smoking, exposure to toxic industrial chemicals and gases, bladder inflammation due to microbial and parasitic infections, as well as some adverse side-effects of medications. The genetic mutations in some chromosomal genes, such as FGFR3, RB1, HRAS, TP53, TSC1, and others, occur which form tumors in the urinary bladder. These genes play an important role in the regulation of cell division which prevents cells from dividing too quickly. The changes in the genes of human chromosome 9 are usually responsible for tumor in bladder cancer, but the genetic mutation of chromosome 22 can also result in bladder cancer. The identification of p53 gene mutation has been studied at NIH, Washington, DC, USA, in urine samples of bladder cancer patients. The invasive bladder cancers were determined for the presence of gene mutations on p53 suppressor gene. The 18 different bladder tumors were evaluated, and 11 (61%) had genetic mutations of p53 gene. The bladder cancer studies have suggested that 70% of bladder cancers involve a specific mutation in a particular gene, namely telomerase reverse transcriptase (TERT) gene. The TERT gene is involved in DNA protection, cellular aging processes, and cancer. The Urothelial carcinomas of the bladder have been described in Atlas of genetics and cytogenetics in oncology and hematology. HRAS is a proto-oncogene and has potential to cause cancer in several organs including the bladder. The TSC1 c. 1907 1908 del (E636fs) mutation in bladder cancer suggests that the location of the mutation is Exon 15 with frequency of TSC1 mutation of 11.7%. The recent findings of BAP1 mutations have shown that it contributes to BRCA pathway alterations in bladder cancer. The discoveries of more gene mutations and new biomarkers and polymerase chain reaction bioassays for gene mutations in bladder cancer need further research.