bluuebunny/crossref_metadata_2025 huggingface dataset:
Sensitivity of tumor cells deficient in the fanconi anemia pathway to inhibition of ataxia telangiectasia mutated (ATM)
Authors: Kennedy R. D. , Stuckert P. , Archila E. , De LaVega M. , Chen C. , Moreau L. , D'Andrea A.
Published: March 2020

10509

Loss of the fanconi anemia (FA) pathway function has been described in a number of sporadic tumor types including breast, ovarian, pancreatic, head and neck and hematological malignancies. Functionally, the FA pathway responds to stalled DNA replication following DNA damage. Given the importance of the FA pathway in the response to DNA damage, we hypothesized that cells deficient in this pathway may become hyper-dependent on alternative DNA damage response pathways in order to respond to endogenous genotoxic stress such as occurs during metabolism. Therefore, targeting these alternative pathways could offer therapeutic strategies in FA pathway deficient tumors. To identify new therapeutic targets we treated FA pathway competent and deficient cells with a DNA damage response siRNA library, that individually knocked out 230 genes. We identified a number of gene targets that were specifically toxic to FA pathway deficient cells, amongst which was the DNA damage response kinase Ataxia Telangiectasia Mutated (ATM). To test the requirement for ATM in FA pathway deficient cells, we interbred Fancg ± Atm± mice. Consistent with the siRNA screen result, Fancg-/- Atm-/- mice were non viable and Fancg± Atm-/- and Fancg-/- Atm ± progeny were less frequent that would have been expected. Several human cell lines with FA gene mutations were observed to have constitutive activation of ATM which was markedly reduced on correction with the appropriate wild-type FA gene. Interestingly, FA pathway deficient cells, including the FANCC mutant and FANCG mutant pancreatic cancer cell lines, were selectively sensitive to monotherapy with the ATM inhibitor KU55933, as measured by dose inhibition and colony count assays. FA pathway deficient cells also demonstrated an increased level of chromosomal breakage, cell cycle arrest and apoptosis following KU55933 treatment when compared to FA pathway corrected cells. We conclude that FA pathway deficient cells have an increased requirement for ATM activation in order to respond to sporadic DNA damage. This offers the possibility that monotherapy with ATM inhibitors could be a therapeutic strategy for tumors that are deficient for the FA pathway.

No significant financial relationships to disclose.
BRCA1/2 mutations and cancer risk in Asian-Americans
Authors: Kurian A. W. , Chun N. M. , Mills M. A. , Staton A. D. , Crawford B. A. , Ridge Y. , Donlon S. S. , Gong G. D. , West D. W. , Ford J. M. ,   Asian-Caucasian BRCA1/2 Study Group
Published: March 2020

10512

Background: There are significant differences in breast cancer epidemiology between Caucasian and Asian-Americans and even between different Asian groups. These cancer risks and associated BRCA1/2 mutation prevalence have not been well defined in Asians; BRCA1/2 mutation penetrance might differ due to different risk modifiers. We report on a case-control study of BRCA1/2 mutation prevalence and cancer risk in Asian-American women. Methods: Clinical Data Collection: Chart review from cancer genetics services of 4 North American centers with highest Asian volume. BRCA1/2 Mutation Risk Assessment: BRCAPRO and Myriad II models, CancerGene version 4.3 (University of Texas). BRCA1/2 Mutation Testing: Full sequencing and large rearrangement panel (Myriad Genetics Inc.). Endpoints: BRCA1/2 mutation prevalence and predictive model accuracy (observed versus predicted mutations). Results: 43 of 181 Asians (23.8%) had a BRCA1/2 mutation; 36 (19.9%) had a variant of uncertain significance. The observed prevalence of BRCA1/2 mutations was 23.8% of women, which differs significantly from the predicted prevalence of 12.9% using BRCAPRO (p = 5.6 × 10-9), and the predicted prevalence of 12.6% using Myriad II. This 2-fold difference existed for Chinese, Japanese, and Filipina women (the ethnic sub-groups with enough cases available for comparisons), even though the percent with observed and predicted mutations varied for these three groups. Conclusions: One in 4 clinically tested Asian-Americans has a BRCA1/2 mutation. Standard models significantly under-predict mutations in Asians; consequently, Asians are likely under-tested for BRCA1/2 mutations. These results may reflect lower BRCA1/2-associated cancer risk in Asians compared to Caucasians. Comparison to Caucasian controls and to Asians in Hong Kong is underway, to investigate potential genetic and lifestyle modifiers of BRCA1/2-associated cancer risk.

[Table: see text]

No significant financial relationships to disclose.
Molecular genetic testing for large genomic deletion and duplication mutations in the BRCA1 and BRCA2 genes for hereditary breast and ovarian cancer
Authors: Wenstrup R. , Judkins T. , Eliason K. , Schoenberger J. , Rajamani S. , Frye C. A. , Burbidge L. A. , Trost J. T. , Deffenbaugh A. M. , Roa B. B.
Published: March 2020

10513

Background: Mutations in the BRCA1 and BRCA2 genes are comprised of a majority of mutations that are detectable by sequence analysis, and a minority of large genomic deletion and duplication mutations that are detected by methods different than sequencing. Our laboratory developed and implemented a clinical assay for large rearrangements that we refer to as BART (BRCA1/2 Rearrangement Test). We validated the performance of BART using a completely anonymous large number of breast/ovarian cancer patient samples. We also demonstrated superior performance of BART versus other dosage-sensitive methods including Multiplex Ligation-dependent Probe Amplification (MLPA). Methods: BART utilizes quantitative endpoint polymerase chain reaction (PCR) in a multiplexed fluorescent format. Eleven multiplex PCR reactions were designed to contain two amplicons targeting the promoter region, all coding exons, and flanking regions of BRCA1 and BRCA2. An automated likelihood-based analysis application normalizes target amplicon copy number between BRCA1, BRCA2 and three control genes. Deletions and duplications are identified with a statistical confidence level. Results: Based on clinical and family history criteria, 1,035 patients were identified as severe-risk during the initial months of clinical BART analysis at Myriad Genetic Laboratories. All patients were initially tested for Comprehensive BRACAnalysis which includes BRCA1 and BRCA2 full gene sequencing plus large rearrangement panel testing for 5 recurrent BRCA1 mutations. Among severe-risk patients, 302 (29.2%) were positive for a BRCA1 or BRCA2 mutation by sequencing, 9 (0.9%) were positive by large rearrangement panel testing and an additional 27(2.6%) tested positive by BART for large genomic rearrangements. The total detection rate for deleterious mutations in severe-risk individuals was therefore 32.7%. Conclusions: Our initial clinical data indicate that BART testing is appropriate for high-risk patients identified on the basis of personal and family history criteria.

No significant financial relationships to disclose.
Increased HER2 expression in women with recurrent ER positive breast cancer
Authors: Kaklamani V. G. , Cicconi J. , Gradishar W. , Willey E. , Diaz L. , Rademaker A. , O'Regan R.
Published: March 2020

10516

Introduction: Tamoxifen (TAM) remains widely used in the treatment of all stages of breast cancer. Although the majority of hormone receptor (HR) positive tumors respond to TAM, many of these breast cancers will develop resistance resulting in disease recurrence or progression. Over-expression of HER2 appears to play a role in de novo tamoxifen-resistance. We have demonstrated previously that HR-positive breast cancers exposed to selective estrogen receptor modulators (SERMs), such as tamoxifen, in vivo continue to express HR but have an increase in the expression of HER2 (O'Regan Clin Cancer Res 2006). However the above finding has not been confirmed in patient samples. Materials and Methods: We evaluated 30 paired tissue samples from patients with HR positive tumors whose cancers recurred. The first tissue sample is from diagnosis and the paired sample comes from metachronous metastatic disease. Results: The median age of diagnosis was 56 (29–96). Seven patients presented with stage I disease, 11 with stage II and nine with stage III3, and three patients had missing staging information. The median time to recurrence was three years. The expression of ER decreased from diagnosis to recurrence from 79% to 59% (p=0.035). PR also decreased between diagnosis and recurrence from 34% to 22% (p=0.13). HER2 score was 2 or 3 in 27% of samples at diagnosis and in 53% at time of recurrence (p=0.01). These cancers did not have HER2 gene amplification. Conclusions: These results confirm our in vivo findings that over-expression of HER2 plays a significant role in acquired TAM- resistance. We have previously demonstrated that trastuzumab inhibits growth of SERM-resistant breast cancers in vivo despite the fact that these cancers did not have HER2 gene amplification. Taken together, our findings suggest that trastuzumab should be examined clinically in patients with TAM-resistant metastatic breast cancer, as they may be particularly sensitive to inhibition of HER2-driven pathways.

No significant financial relationships to disclose.
p63/p73 expression mediates cisplatin sensitivity in a subset of triple-negative primary breast cancer: Implications for a new clinical trial
Authors: Isakoff S. J. , Leong C. , Vidnovic N. , DeYoung M. P. , Sgroi D. , Goss P. E. , Ryan P. D. , Ellisen L. W.
Published: March 2020

10522

Background: Triple-negative breast cancer (ER-, PgR- and HER2-negative) has a poor prognosis relative to other subtypes, even with the best available chemotherapy. We recently reported that in squamous cell cancers, p63 promotes survival by direct interaction with and inhibition of p73-dependent apoptosis. We now report on the role of p63/p73 in triple-negative breast cancer. Methods: We performed quantitative RT-PCR on primary breast cancer specimens to determine the expression of specific p63 and p73 isoforms. Cell culture experiments used MCF7, HCC-1937, MDA-MB-468, T47D, and MCF10A cell lines. Lentiviral p63 and p73 shRNA were used for knockdown experiments. Chemotherapy dose-response curves were generated using MTT assay. Results: In primary tumors, ΔNp63 and TAp73 isoforms are expressed exclusively within a subset of triple-negative tumors in which p53 mutations are commonly found. Knockdown of p63 in triple-negative cell lines, but not the ER-positive MCF7 line, induced apoptosis which was specifically dependent on p73 expression. Breast cancer cell lines expressing p63/p73 exhibited p73-dependent sensitivity to cisplatin. Knockdown of p73 resulted in a 10–100 fold increase in the IC50 of cisplatin, but no change in doxorubicin or paclitaxel sensitivity. Exogenous p73 expression in MCF10A cells, which have high levels of p63 but no p73, confers increased sensitivity to cisplatin. Conclusion: These results show that p63/p73expression promotes survival in a subset of breast cancers, and they provide a mechanism for cisplatin sensitivity. These findings suggest that p63/p73 expression may serve as a biomarker to predict cisplatin sensitivity in triple-negative breast cancer. Based on these results, we are conducting a phase II study of 39 patients evaluating cisplatin as first line therapy for metastatic triple-negative breast cancer. The primary endpoints are overall objective response rate and response rate in low versus high p63/p73 expressing subgroups.

No significant financial relationships to disclose.
FLT3 D835/I836 mutations predict worse disease-free survival (DFS) in younger adults with cytogenetically normal acute myeloid leukemia (CN AML) without FLT3 internal tandem duplications (ITD): A Cancer and Leukemia Group B (CALGB) study
Authors: Whitman S. P. , Ruppert A. S. , Radmacher M. D. , Mrozek K. , Paschka P. , Kolitz J. E. , Larson R. A. , Caligiuri M. A. , Marcucci G. , Bloomfield C. D.
Published: March 2020

10532

Background: The adverse prognostic impact of FLT3 ITD is well-recognized in younger adults with CN AML, whereas the clinical relevance of D835/I836 mutations in the second tyrosine kinase domain of FLT3 (FLT3 TKD), that occur in 5%-14% of patients (pts), remains controversial. We evaluated the prognostic impact of FLT3 TKD, alone and with other prognostic markers, in 139 de novo CN AML adults, aged <60 years, without FLT3 ITD, and treated on similar CALGB protocols (9621 and 19808). Methods: FLT3 TKD was detected by RT-PCR/EcoRV digestion. Affymetrix U133 plus 2.0 gene expression profiles derived from FLT3 TKD+ and FLT3 wild-type (FLT3 WT) pts with available RNA were compared. Results: Relative to FLT3 WT pts, FLT3 TKD+ pts [n=16 (11.5%)] presented with higher white blood cell counts (WBC) (median: 57.2 v 14.6 × 109/L; P<.001), % bone marrow (BM) blasts (median: 75.5 v 63; P=.001), and % blood blasts (median: 62.5 v 46; P=.03). All 16 FLT3 TKD+ pts achieved complete remission versus 85% of FLT3 WT pts (P=.13). With a median follow-up of 4.6 years, FLT3 TKD+ pts had worse DFS (P=.01; 3-year DFS rates: 31% v 60%) and a trend for worse overall survival (OS) (P=.17, 3-year OS rates: 40% v 65%) compared to FLT3 WT pts. Other variables moderately associated with a worse DFS (P<.10) included higher % BM blasts and NPM1 WT. Adjusting for these two variables, FLT3 TKD remained significantly associated with adverse DFS (P=.02; hazard ratio=2.3). A gene expression signature consisting of 333 probe sets (P<.001) differentiated FLT3 TKD+ (n=9) from FLT3 WT (n=50) pts. Genes represented in this signature include those involved in cell motility (e.g., C3AR1, VNN1, VNN2, ADAM8; all over-expressed in FLT3 TKD+), development (e.g., HOXB5, HOXB6; both over-expressed in FLT3 TKD+) and stress response (e.g., MPO; under-expressed in FLT3 TKD+). Conclusions: FLT3 TKD appears to be an independent predictor for DFS among younger CN AML adult pts without FLT3 ITD. Gene expression profiling provides additional insight into the biology of FLT3 TKD+ AML and may lead to identification of additional targets for improved therapy.

No significant financial relationships to disclose.
Clinical implications of the molecular relationship between HER2 and BRCA1
Authors: Ellsworth R. E. , Deyarmin B. , Hooke J. A. , Shriver C. D.
Published: March 2020

10536

Background: While HER2 status is used prognostically, the effects of HER2 can be influenced by other genes; co- amplification of TOP2A modulates the effectiveness of treatment with Herceptin. BRCA1, deleted in >50% of sporadic breast tumors, is located distal to HER2 on chromosome 17q and is marked by structural instability. To determine how alterations at the BRCA1 locus may modify amplification of HER2, FISH analysis of BRCA1 was performed in breast tumors with HER2 amplification and/or allelic imbalance on chromosome 17q12. Methods: HER2 status (n=68) was determined using the PathVysion HER2 kit. For BRCA1 analysis, DNA probes were generated by nick translation from a BAC clone containing the entire BRCA1 gene in conjunction with a CEP 17 probe. Copy number gains and deletions were defined as BRCA1:CEP17 >1.3 and <0.8, respectively; aneusomy was defined when >10% of tumor cells had copy numbers > (polysomy) or < (monosomy) than two. Results: Amplification of the HER2 gene was detected in 57% of tumors; an additional 16% of tumor were polysomic for chromosome 17q. The remaining 27% of tumors had previously detected allelic imbalance at the 17q12 region. The majority of samples (47%) were characterized by an amplification of HER2 with a downstream deletion of BRCA1, while HER2 and BRCA1 were amplified concomitantly in 21% of tumors, including those with polysomy of 17q. Fifteen percent of samples were monosomic for chromosome 17q; using current clinical scoring criteria, each of those samples were reported as having, despite the altered copy number, negative HER2 status. Levels of overall genomic instability, which were previously measured at 26 chromosomal regions, were significantly higher (P<0.05) in tumors with deletion of BRCA1 compared to those without loss of BRCA1 (38% and 24%, respectively). Conclusions: Alterations, especially deletions, of BRCA1 are highly correlated with copy number changes in HER2. Because deletion of BRCA1 has been associated with overall genomic instability, patients with amplification of HER2 and deletion of BRCA1 may prove unresponsive treatment with Herceptin. Loss of BRCA1 should, therefore, be considered as a genetic modifier to HER2 amplification and the status of BRCA1 should be considered in selecting treatment options for patients with HER2 amplified tumors.

No significant financial relationships to disclose.
Genotype and phenotype in hereditary and sporadic breast cancers
Authors: Giorgetti G. , Galizia E. , Bianchi F. , Ferretti C. , Corradini F. , Belvederesi L. , Piccinini G. , Loretelli C. , Santinelli A. , Cellerino R.
Published: March 2020

10538

Background: BRCA1 protein is involved in distinct DNA-repair processes. Germline mutations in BRCA1 gene confer cancer susceptibility. A frequent mechanism for epigenetic inactivation is hypermethylation of the CpG island in promoters of tumours suppressor genes. BRCA1 promoter hypermethylation has been found in a variable percentage of breast cancers (15–30%). BRCA1-associated breast cancers are usually high-grade, poorly differentiated and stain negative for HER2/neu, oestrogen and progesterone receptors (ER, PgR). Many studies have shown that hereditary BRCA1 and basal-like sporadic breast tumours have a similar phenotype and gene expression signature. Methods: By clinical criteria, 223 patients were selected and, for each patient, the probability to carry a BRCA1 mutation was calculated using the software BRCAPRO and Manchester Score System. All patients were studied by direct sequencing and MLPA of BRCA1 Open Reading Frames (ORFs). Thirty sporadic breast carcinomas, from women undergone surgery for primary invasive breast carcinoma between 1995 and 2001, were selected on the basis of negative staining for ER, PgR and HER2/neu (“BRCA-like”). In these patients, Methylation Specific-PCR and Bisulfite Sequencing on genomic DNA (obtained from sections of paraffin-embedded tissues and modified with sodium bisulfite) were used to assess the methylation pattern of BRCA1 promoter. BRCA1 immunohystochemical analysis (IHC) was performed in all patients. Results: We identified 17 patients with deleterious germline mutations in BRCA1. In “BRCA-like” patients, 13 methylated and 17 unmethylated cases were found by methylation analysis of BRCA1 promoter. The BRCA1 IHC was performed in all available samples ( table 1 ). Conclusions: Hypermethylation of BRCA1 promoter was found in 43% of “BRCA- like” patients. Expression of BRCA1 seems to correlate with hypermethylation of its promoter. Further studies are in progress to better understand the possible role of BRCA1 promoter hypermethylation in sporadic breast cancers.

[Table: see text]

No significant financial relationships to disclose.
Screening an Irish cohort with colorectal cancer for Lynch Syndrome using immunohistochemistry for mismatch repair proteins
Authors: Power D. G. , Farrell M. P. , Muldoon C. B. , Fitzpatrick E. , Stuart C. , Flannery D. , Kennedy M. J. , Stephens R. B. , Daly P. A.
Published: March 2020

10547

Background: Large-scale screening for germ-line mutations that lead to the onset of disease in adulthood is possible owing to recent technical advances. The care of those with inherited predisposition to breast and ovarian cancer is now becoming a mainstream component of medical care. It is more difficult to identify those with Lynch Syndrome (LS) as various criteria (Amsterdam and Bethesda) have not proved definitive. An important development is the examination of tumor tissue to detect mismatch repair (MMR) protein loss using immunohistochemical (IHC) techniques. When coupled with family history those at risk of harbouring a mutation for LS can be identified. Once a mutation is identified predictive testing can be offered to family members, risk-reduction measures applied and mortality from colorectal cancer reduced. Methods: Screening for MMR protein expression (MLH1, MSH2, MSH6, PMS2) was planned on all colorectal cancer (CRC) cases using IHC on formalin-fixed tumor tissue from January 1st 2002. Local ethics committee approval was obtained and then written informed-consent from patients. Family history data was gathered from the index case or an appropriate relative. An aliquot of blood was stored from index cases for subsequent genetic screening if indicated by IHC analysis and genetic counseling. Results: 108 cases with CRC (62 male, 46 female, median age 59 years) from a potential total of 612 have been screened for MMR protein expression by a gastrointestinal pathologist and independently validated. Turn-around time for IHC analysis was 9 weeks. 5 patients (4.6%) had loss of MMR proteins, MSH2/MSH6- 2 cases, MSH6 alone- 1 case and MLH1/PMS2- 2 cases. All 5 have opted for genetic counselling and sequencing of relevant genes. Conclusion: These early results in an Irish cohort with CRC showing MMR loss in 4–5% of cases is consistent with other population findings. Microsatellite instability analysis is difficult, expensive and relatively unavailable. IHC, however, is an established technique in pathology departments and can be the cheapest and most reproducible approach to identify LS cases. IHC results along with robust family data can guide the genetic counseling process towards preventing deaths from CRC and other LS-associated cancers.

No significant financial relationships to disclose.
The 8.1 ancestral MHC haplotype is strongly associatied with colorectal cancer
Authors: Kocsis J. , Toth K. E. , Madaras B. , Biro A. , Füst G. , Blaskó B. , Pocsai Z. , Ádány R. , Laki J.
Published: March 2020

10548

Background: The immune system is believed to play an important role in sufficiently or insufficiently inhibiting the multistep process of tumour development. The so-called 8.1 ancestral haplotype (8.1AH) in the main histocompatibility complex is associated with alterations of the immune response characterized by an increase in antibodies, circulating immune complexes and TNF-a, and lack of a complement protein component (C4A). Methods: To study the possible association of the 8.1AH with colorectal cancer the DNA samples of 183 hungarian patients (average age 67.99 ys; 63–74 ys) with colorectal cancer and 129 healthy hungarian controls (average age 66.5 ys; 60–73.5 ys) were genotyped for member alleles of the 8.1 AH: AGER -429C, HSP70–2 -1267G (HSP70–2G), TNF- a -308A (TNF2) and LTA+252G by SSP-PCR and PCR-RFLP. Simultaneous carriage of the four member alleles was considered carriage of the 8.1AH. Results: We found the frequency of the 8.1AH to be significantly higher in patients with colorectal cancer than in healthy controls (19.1% vs. 7%, P=0.002). After adjustment to age and gender individuals carrying the 8.1AH had a 2.81 (1.23–6.22, P=0.011) fold risk to develop colorectal cancer compared with non-carriers of the haplotype. None of the member alleles alone showed association with colorectal cancer (PAGER-429C=0.486; PHSP70–2G=0.898; PTNF2=0.140; PLTA=0.376.) Conclusions: The 8.1AH by the genetically encoded alterations of the immune response may affect the development of colorectal cancer. The frequency of the 8.1AH is 5–10% in Hungary/Central Europe but, for instance, approx. 20–25% in the populations with Celtic origin. Our novel observation might contribute to cancer research and therapy when more information is available on which key components of the altered immune response encoded by the 8.1AH are not effective enough against tumor development.

No significant financial relationships to disclose.
Breast cancer survival in carriers of BRCA1 and BRCA2 mutations
Authors: Rennert G. , Bisland S. , Bar Joseph N. , Zhang S. , Rennert H. S. , Barnett-Griness O. , Narod S. A.
Published: March 2020

10552

Background: The characteristic pathologic features of breast cancers in women with a BRCA1 mutation suggest that women with hereditary breast cancer might have a worse than expected prognosis, but the results of clinical studies have been inconsistent. The current study is a national population-based study of Israeli women aimed at evaluating the impact of the inheritance of a BRCA1 or a BRCA2 mutation on breast cancer prognosis. Methods: All incident cases of invasive breast cancer diagnosed in Israel from January 1, 1987 to December 31, 1988 were identified. Paraffin-embedded tumor blocks or unstained slides and pathology and oncology records were requested for all patients. DNA was extracted from each paraffin block and was analyzed for three founder Jewish mutations in BRCA1 and BRCA2. Of 2,514 diagnosed cases, a pathology sample was retrieved from 1,794 (71.4%) and the medical record was retrieved for 1,545 of these (86.1%). Results: A BRCA1 or BRCA2 mutation was identified in 10% of Ashkenazi women with breast cancer, including 18% of women diagnosed below the age of 50. The adjusted hazard ratio for breast cancer-specific survival was not different for non-carriers and carriers of a BRCA1 (HR = 0.8, 95% CI: 0.5 - 1.3, p = 0.3) or BRCA2 (HR = 1.3, 95% CI: 0.8 - 2.2, p = 0.3) mutation. Among women who were treated with CMF/CAF chemotherapy, BRCA1 carriers experienced better survival than non-carriers (HR = 0.5; 95% CI 0.2–1.2) but the difference did not reach statistical significance (p = 0.1). Tumor size (>2 cm) was a significant predictor of breast cancer specific mortality in non-carriers (HR = 2.8; 95%CI: 2.2 to 3.5) but not in BRCA1 carriers (HR = 1.1; 95% CI 0.5 to 2.8). ER status also did not play a significant predictive role in survival of carriers. Conclusions: Compared to non-carriers, women with breast cancer who carry a founder Jewish mutation in one of the BRCA genes experience similar or possibly even lower breast cancer-specific mortality rate, in spite of a bad profile of prognostic factors.

No significant financial relationships to disclose.
MMP1 1G/2G promoter polymorphism and risk of esophageal adenocarcinoma
Authors: Singh S. , Asomaning K. , Kulke M. H. , Zhou W. , Zhai R. , Su L. , Heist R. S. , Lynch T. J. , Christiani D. C. , Liu G.
Published: March 2020

10555

Background: The 2G allele of the MMP1 -1607 1G/2G promoter polymorphism creates an Ets binding site that leads to increased transcriptional and enzyme activity, particularly in the presence of growth factors and cytokines. This polymorphism has been associated with greater risk of cancer (e.g., renal cell, lung and oral cancers, glioblastomas) and cancer invasiveness (e.g., melanoma, cervical, lung and colorectal cancer). The aim of the current study was to evaluate the role of this MMP 1G/2G polymorphism in the risk of esophageal adenocarcinoma (EA). Methods: We evaluated 323 histologically confirmed EA cases and 464 healthy controls frequency-matched for age and gender. Genotyping of the MMP1 1G/2G promoter polymorphism involved a Taqman approach. All EAs had endoscopic evidence showing that the center of the tumors were located at or above the gastroesophageal junction. Odds Ratios (OR) were calculated using multivariate logistic regression, adjusted for age, gender, smoking status, and body-mass index (BMI) at the age of 18 years (to represent a healthy adult BMI). Results: Genotype frequencies were: 33% (1G/1G), 47% (1G/2G) & 20% (2G/2G) in controls; in cases, 26% (1G/1G), 50% (1G/2G) & 24% (2G/2G). 88% of cases were male. The MMP1 2G/2G and 1G/2G genotypes conferred a greater risk of EA, with adjusted ORs of 1.50 (95%CI=1.0–2.3) and 1.34 (95%CI=0.9–1.9), respectively, when compared with the wildtype 1G/1G genotype. The 2G allele (2G/2G + 1G/2G) conferred an adjusted OR of 1.38 (95%CI=1.0–1.9). By stage, the adjusted ORs for the 2G allele were 1.26 (95%CI=0.8–2.1), 1.45 (95%CI=0.9–2.3), & 1.54 (95%CI=0.9–2.7) for node negative, node-positive, and metastatic disease, respectively. Conclusions: The 2G allele of the MMP1 -1607 1G/2G polymorphism was associated with an increased risk of EA in this analysis. In addition, there was a non-significant trend towards conferring greater risk in the more advanced stages of EA, suggesting a possible role of this polymorphism in the invasiveness of this cancer.

No significant financial relationships to disclose.
Application of serum proteomic patterns by ProteinChip SELDI system in diagnosis of gastric cancer
Authors: Lim J. , Cho J. , Paik Y. , Chang Y. , Kim H.
Published: March 2020

10559

Background: Gastric cancer is one of the most common malignancy in the world and one of the leading causes of cancer related death in Korea. Most treatments for advanced gastric cancer have limited efficacy. So early detection of gastric cancer could have profound impact on the successful treatment. Application of multiple biomarkers may improve the diagnostic prediction to distinguish cancer from non-cancer. ProteinChip Surface-Enhanced Laser Desorption/Ionization Time-of-flight Mass Spectrometry (SELDI-TOF-MS) system is one of the currently used techniques to identify biomarkers for cancers. In this study, we have explored whether the serum proteomic patterns by ProteinChip SELDI system can differentiate gastric cancers from non-cancer cohorts. Methods: We have screened protein profiles of 100 serum samples obtained from 60 gastric cancer patients and 40 healthy individuals. Protein expression profiles were expressed on ProteinChip Array and analyzed by PreoteinChip Reader. Peak intensities were normalized by total ion currency and analyzed by the Biomarker Wizard Software to identify the peaks showing significantly different intensities between normal and cancer groups. Classification analysis and construction of decision trees were done with the Biomarker Pattern Software. Results: SELDI -TOF-MS by averaging 50 laser spots collected at a laser intensity setting of 160, a detector sensitivity of 6, and mean mass range of 30 kDa. Seventeen protein peaks shown significant differences between two groups were chosen to make a protein biomarker pattern. The decision tree which gives the highest discrimination included four peaks at 5,919, 8,583, 10,286, and 13,758 as splitters. The sensitivity and the specificity for classification of with the decision tree giving the highest discrimination were 96.7% (58/60) and 97.5% (39/40), respectively. When the protein biomarker pattern was tested with the blinded test set including 30 gastric cancer patients and 20 healthy individuals and, it yielded a sensitivity of 93.3% (28/30) and a specificity of 90% (18/20). Conclusions: These results suggest that serum-protein profiling pattern by SELDI system may distinguish gastric cancer patients from normal counterparts with relatively high sensitivity and specificity.

No significant financial relationships to disclose.
Quantitative immunohistochemical detection of gamma-H2AX in paraffin-embedded human tumor samples at National Clinical Target Validation Laboratory
Authors: Yang S. X. , Nguyen D. , Steinberg S. M. , Ji J. , Parchment R. , Kinders R. , Kummar S. , Gutierrez M. , Murgo A. , Tomaszewski J. E. , Doroshow J. H.
Published: March 2020

10565

Background: DNA double-strand breaks (DSBs) caused by exposure to DNA damaging agents initiate phosphorylation of histone H2AX to form gamma-H2AX, which is considered a surrogate marker of DSBs. However, it is a challenge to quantitatively measure gamma-H2AX in clinical samples such as tumor biopsies. The aim of this study was to develop an immunohistochemical detection method for gamma-H2AX and to quantitatively evaluate the levels of gamma-H2AX in paraffin-embedded tumor samples. Methods: Human breast cancer MCF-7 cells were treated with the topoisomerase I inhibitor irinotecan at 1 μM or vehicle for 1 h, and fixed in 10% neutral buffered formalin and embedded in paraffin. Staining with gamma-H2AX antibody was performed on sections of treated MCF-7 cells, tumor specimens, and biopsies at baseline and after doxorubicin-containing chemotherapy from cancer patients. Numbers of foci and level of gamma-H2AX expression per tumor nucleus were determined by manual counting under a light microscope and an Automated Cellular Imaging System. Results: There was a rise in the mean numbers of nuclear foci and intensity of gamma-H2AX in MCF-7 cells treated with irinotecan versus vehicle (19.9 ± 2.7 vs. 9.95 ± 3.6; P < 0.0001 and 61.2 ± 8.5 vs. 16.2 ± 13.6; P < 0.0001 by Wilcoxon rank sum test). The level of gamma-H2AX foci in human tumor samples was 18.8 ± 13.1, 44.8 ± 14.5, 51.2 ± 20.8, or 69.7 ± 21.2 in carcinomas of the breast, colon, ovary, or prostate. In a patient with stable disease, levels of gamma-H2AX foci were 62.7 ± 26.9 at baseline and 67.2 ± 25.3 after doxorubicin-containing regimen chemotherapy. Conclusions: Our data suggest that the quantitative immunohistochemical detection of gamma-H2AX levels is facilitated by a digital imaging system, and is a reliable method to measure the effects of DNA damaging agents in cells and paraffin-embedded human tumor samples. Its application may help evaluate tumor response to various DNA damaging agents currently in the clinic and those presently undergoing clinical development.

No significant financial relationships to disclose.
Elevated phosphorylated mammalian target of rapamycin (p-mtor) expression in human neoplasms
Authors: Ross J. S. , Qian J. , Sheehan C. , Jones D. , Jennings T.
Published: March 2020

10571

Background: Mammalian target of rapamycin (mTOR) regulates several cell functions critical to tumorigenesis including proliferation, growth, mobility and survival. mTOR expression and dysregulation have been reported carcinomas of breast, head & neck, liver and kidney, but have not yet been studied in non small cell lung cancers (NSCLC), colorectal adenocarcinomas (CRC), nor brain tumors including gemistocytic astrocytomas (GA) and glioblastomas (GBM). Methods: Formalin-fixed paraffin-embedded tissue sections from 121 NSCLC [45 squamous cell carcinomas (SCC), 45 pure adenocarcinomas (AC), and 31 bronchioloalveolar carcinomas (BAC) including both pure BAC and adenocarcinomas with BAC features], 108 CRC, 8 GA and 24 GBM were immunostained by an automated method (Ventana Medical Systems, Tucson, AZ) using monoclonal rabbit anti-human p-mTOR antibody (Cell Signaling, Danvers, MA). Immunohistochemical assessment included intensity and percentage of positive cells in both benign and carcinomatous elements. Results were correlated with morphologic and prognostic variables in NSCLC and CRC and GBM. Results: Elevated p-mTOR expression was noted in 33% SCCs, 64% ACs, and 65% BAC, 78% CRC, 75% GA and 96% GBM. For NSCLC, increased expression of p-mTOR correlated with histologic subtype (p=0.004). Within SCC greater than 3 cm in size, only 20% had an increase in p-mTOR, while 53% of SCC ≤ 3 cm showed an increase in p-mTOR expression (p=0.03). 100% high grade vs. 43% low grade (p=0.05) and 100% advanced stage (p=0.037) vs. 54% low stage BAC showed an increase in p-mTOR. Consistently, 100% BACs with positive lymph node status vs. 54% node negative BACs showed an increased level of p-mTOR (p=0.037). For CRC, p-mTOR over-expression correlated with high tumor grade (80% Grade 3 vs 50% Grade 1 p=0.042). Majority of GA (6/8) and nearly all GBM (23/24) were positive for p-mTOR (p=0.135). Conclusions: Activated mTOR protein, defined by phosphorylation of mTOR at Serine-2448 (p-mTOR), is increased in NSCLC, CRC and both GA and GBM. These findings suggest a potential role for mTOR dysregulation in lung, colon and brain tumorigenesis and support the currently ongoing clinical trials investigating the therapeutic relevance of rapamycin analogs in the treatment of these tumors.

No significant financial relationships to disclose.
Targeting the mitotic spindle in the treatment of adrenocortical cancer
Authors: Stephan E. A. , Gonzales P. , Han H. , Grant C. S. , Gately S. , Von Hoff D. D. , Trent J. M. , Demeure M. J.
Published: March 2020

10572

Background: Adrenocortical carcinoma (ACC) is a rare and lethal cancer with an average incidence of 1 in 1.4 million. Response to current standard therapy including mitotane is approximately 22%. Herein we use gene expression profiling to identify novel targets for therapeutic intervention. Methods: The transcriptomes of 29 ACC tumors and 4 normal adrenal glands were profiled on Affymetrix Human Genome and Agilent 22K element expression arrays in order to identify genes that could represent potential therapeutic targets. Agents known to inhibit the selected targets were tested in comparison with mitotane in two ACC cell lines (H295R and SW-13) in vitro and are currently being tested in a Scid mouse xenograft system. Results: Expression of numerous kinesin family members is dysregulated within the ACC tumor population. Of interest in the ACC cells EG5 (KIF11, KSP), a kinesin important for mitotic spindle formation, is up-regulated 5.81+ 4.42 (μ+SD) and 6.48 + 0.99 fold on the Agilent and Affymetrix platforms respectively. In addition, SPARC, an albumin binding matrix associated protein is up-regulated in ACC tumors by 3.69 + 2.37 (μ+SD) and 1.56 + 0.44 on the two platforms. The anti-microtubular drugs paclitaxel and albumin bound paclitaxel (abraxane) exhibited in vitro inhibition of H295R and SW-13 cells at IC50 concentrations of 0.33 μM and 0.0078 μM for paclitaxel and 0.35 μM and 0.0087 μM for Abraxane compared to mitotane concentrations of 15.9 μM and 46.4 μM. Treatment effects of monastrol, paclitaxel, and Abraxane are currently being tested in xenograft murine models. Testing of the EG5 inhibitor S-trityl-L- cysteine is ongoing in the in vitro system. Conclusions: KIF11 inhibitors have been proposed in the treatment of cancer. These compounds may offer better anti-mitotic activity than other tubulin interacting agents with less neurotoxic side effects. Based on biological insight garnered through expression profiling of ACC tumors, the current ongoing research is investigating the use of abraxane and EG5 inhibitors in the in vitro and in vivo treatment of ACC.

No significant financial relationships to disclose.
Rethinking the role of nodal status in TNM staging of node-positive breast cancer
Authors: Anderson S. J. , Bryant J. L. , Jeong J. , Tang G. , Costantino J. P. , Mamounas E. , Geyer C. E. , Wolmark N.
Published: March 2020

10584

Background: Among TNM staging criteria, the number of positive lymph nodes [PNOD] is the strongest predictor of patient outcome. Based on SEER data, Vinh Hung (2004) asserted that the ratio of the number of positive nodes to the number of nodes removed [NODR] may better predict outcome. We evaluated this assertion in patients from 13 node-positive NSABP phase III trials. Methods: The impact of PNOD, its logarithm (LPNOD) and NODR on survival (S) and recurrence free survival (RFS) were assessed via Cox PH models in 19,768 patients with node-positive breast cancer in 13 NSABP clinical trials. The cohort was split into 2 groups: 9 older trials (Protocols B-05, B-07, B-08, B-09, B-10, B-11, B-12, B-15 and B-16) consisting of 9,915 patients and 4 newer trials (B-22, B-25, B-28 and B-31) consisting of 9,853 patients. Only randomized eligible patients with follow-up were considered. Models were fit to the outcomes for the older trials and validated in the newer trials. Results were adjusted for age, protocol and tumor size. We used Wald z-scores, Nagelkerke R2, and residual analyses to assess the strength of the relationship between nodal status and outcomes. Results: Median time on study was 19.8 years in the older trials and 10.6 years in the newer trials. Ten-year models across the protocols revealed that the relative hazard ratio for each of the outcomes studies was logarithmic according to the number of positive nodes for = 30 positive nodes. Using LPNOD and NODR compared to PNOD improved the strength of association but did not greatly improve goodness of fit ( Table 1 1). For all endpoints, the strength of association and goodness of fit associated with nodal status and other TNM factors attenuated in newer protocols. Conclusions: Nodal status and other TNM factors currently used for breast cancer staging continue to be strong indicators of patient prognosis. However, these factors should be augmented to account for new genetic and biological markers.

[Table: see text]

No significant financial relationships to disclose.
CDK2 and CDK1 specific activities: A novel prognostic indicator in early breast cancer
Authors: Kim S. , Masuda N. , Inaji H. , Yoshidome K. , Tsujimoto M. , Akiyama F. , Ishihara H. , Hortobagyi G. N. , Ueno N. T. , Noguchi S.
Published: March 2020

10592

Background: Cyclin-dependent kinases (CDKs) are expressed almost constantly but their activities change according to cell cycle phase. We studied the specific activity (SA; activity/expression) of CDKs to accurately evaluate their role in cell proliferation. We focused on the ratio of CDK2 SA to CDK1 SA because this ratio has been associated with rapid tumor growth in human breast cancer xenografts. Our goal was to investigate the prognostic significance of CDK2/1 ratio in node-negative breast cancer. Methods: We used a novel assay to measure SAs of CDK2 and CDK1 in 365 primary breast tumors. All tumors were histologically confirmed invasive breast cancer without lymph node involvement. The primary endpoint was relapse-free survival (RFS). Cut-off values for CDK2/1 ratio were defined as those points that best discriminated groups according to RFS. Tumors were grouped as low (136pts), intermediate (84pts), and high (145pts) CDK2/1 ratio. The median follow-up was 59 months (range 4–102). Results: Pts characteristics are described as follows: menopausal status: pre- 42%, post- 58%; tumor size: =20mm 48%, >20mm 52%; histologic grade (HG): I 28%, II 47%, III 23%; ER: (+) 57%, (-) 42%; PR: (+) 51%, (-) 48%. Pts with HG III or high CDK2/1 ratio showed significantly lower 5y-RFS rates than those with HG I/II or low/intermediate CDK2/1 ratio, respectively (HG: I 97%, II 91%, III 85%, P=0.040; CDK2/1 ratio: low 97%, intermediate 92%, high 85%, P=0.017). In univariate analysis, PR (- vs +, P=0.088) had a tendency to associate with relapse, and HG (III vs I, P=0.024) and CDK2/1 ratio (high vs low, P=0.011) also had a significant correlation with relapse. However, only CDK2/1 ratio showed a significant independent prognostic indicator in multivariate analysis (hazard ratio 2.86, 95% CI 1.25–6.58, P=0.01). More important, among women (n=185) with hormone receptor (HR) positive disease given adjuvant hormone therapy alone, who have had no useful factors to predict their outcomes, high CDK2/1 ratio was also associated with worse prognosis than low CDK2/1 ratio (5y-RFS 84% vs 100%, P=0.007). Conclusions: For patients with node- negative disease, especially those with HR positive tumors given adjuvant hormone therapy alone, the CDK2/1 ratio might be useful as a routine laboratory test to predict outcome.

No significant financial relationships to disclose.
Predictive value for survival of a risk model of two serological markers, beta-2-microglobulin (B2M) and lactic dehydrogenase (LDH), in diffuse large cell lymphoma (DLCL)
Authors: Rodriguez M. A. , Temple S. , Fayad L. , Hagemeister F. , McLaughlin P. , Romaguera J. , Cabanillas F.
Published: March 2020

10599

Background: The International Prognostic Index (IPI), the standard tumor risk model in patients (pts) with diffuse large cell lymphoma (DLCL), has 5 factors (age, LDH, performance status, extra nodal sites, and stage). We previously proposed a simpler two-factor model, based on LDH and B2M (JCO10;1989). B2M is a component of HLA class I antigens, expressed in lymphocytes, and a known prognostic indicator in some lymphoid malignancies. The benefits of this model are: objective measures; lab method widely available; simplicity; biologic marker. Methods: We applied the model to a large cohort of DLCL patients with prospective baseline B2M, and treated with doxorubicin-based (chemo) regimens, with and without rituximab. 718 pts with DLCL were sequentially treated at MDACC by IRB approved chemo protocols from 1988–2000. In 2001, rituximab plus chemo (RCHOP) became standard. 311 DLCL pts were sequentially treated with RCHOP from 2001–2005. Cox regression analyses for univariate and multivariate models of IPI factors and B2M were done. Kaplan-Meier survival projections were in three risk categories: low (normal [nl] LDH and B2M); intermediate (either LDH or B2M > nl); or high (LDH and B2M > nl). Results: In both treatment groups, IPI factors and B2M were significant as univariate factors. In the RCHOP group, however, the IPI multivariate model showed age, stage, and extra-nodal sites were not significant risk factors, while B2M and LDH remained highly significant (p<0.01). The 5 year survival projections by risk category were: (*) combines intermediate low and intermediate-high categories Conclusions: This simple two-factor model predicts risk for patients with DLCL, treated with or without rituximab, comparably to the IPI. B2M should be considered an important prognostic indicator in DLCL, particularly in rituximab treated patients. Exploratory analyses to revise the IPI model are indicated.

[Table: see text]

No significant financial relationships to disclose.
Clinical significance of Cdh1-Skp2 cascade in breast cancer
Authors: Fujita T. , Wu G. , Wood R. D. , Wan Y.
Published: March 2020

10609

Background: Skp2 (S-phase kinase-associated protein 2) is substrate-recognizing subunit of an ubiquitin E3 ligase SCF (Skp1, Cullin and F-box) complex and has a major role in p27 regulation. Overexpression of Skp2 is frequently seen in human tumors, often correlating with poor prognosis. Anaphase-promoting complex/cyclosome (APC/C) is another most prominent ubiquitin E3 ligase in cell cycle control, and its function is tightly controlled by activating subunit Cdh1 (Fizzy-related). APC-Cdh1 complex is supposed to be involved in Skp2 proteolysis and have essential role for S phase entry, however clinical significance of Cdh1 have not evaluated yet. Herein we examined the significance of Cdh1 in breast cancer. Methods: Firstly, MCF7 human breast cancer cells and MCF10A normal breast epithelial cells were analyzed in vitro and in vivo. Thereafter, using tissue microarray, we evaluate the expression profile of Cdh1, Skp2 and p27 in both breast cancer and normal breast epithelial tissue. Moreover, clinicopathological significance (age, tumor size, lymph node metastasis, distant metastasis, histological grade, and stage) of Cdh1 was analyzed from 126 breast cancer patients. Chi square-test, Fisher's exact test were used for statistical analysis of immunostaining results and clinicopathological data. p<0.05 was considered statistically significant. Results: Overexpression of Cdh1 induced attenuation of Skp2 and increased p27 protein expression, resulted in growth suppression. Moreover, knockdown of Cdh1 promoted higher Skp2 expression and S-phase population, reduced p27 and consequently induced cell transformation and proliferation in vitro and in vivo. Tissue microarray results appeared that positive Cdh1 and p27 was more frequently seen in normal breast tissue and statistically significant. On the other hand, Skp2 was less in normal tissue. Furthermore, Cdh1 positive breast cancer was more frequently seen in low histological grade tumors and statistically significant (p=0.04). Conclusions: Skp2 protein expression is regulated by Cdh1 in breast cancer. Cdh1 expression could be possible novel biomarker in patient with breast cancer.

No significant financial relationships to disclose.