Druggable Genetic Alterations Research Articles

Determinants of immunological evasion and immunocheckpoint inhibition response in non-small cell lung cancer: the genetic front.

The incorporation into clinical practice of immune-checkpoint inhibitors (ICIs), such as those targeting the cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) and the programmed cell death 1 (PD-1) and its ligand (PD-L1), has represented a major breakthrough in non-small cell lung cancer (NSCLC) treatment, especially in cases where the cancer has no druggable genetic alterations. Despite becoming the standard of care in certain clinical settings, either alone or in combination with chemotherapy, a proportion of patients do not respond while others actually progress during treatment. Therefore, there is a clinical need to identify accurate predictive biomarkers and to develop novel therapeutic strategies based on ICIs. Although they have limitations, the current markers evaluated to select which patients will undergo ICI treatment are the levels of PD-L1 and the tumor mutational burden. In this paper we describe what is currently known about the dynamic interaction between the cancer cell and the immune system during carcinogenesis, with a particular focus on the description of the functions and gene alterations that preclude the host immunoresponse in NSCLC. We emphasize the deleterious gene alterations in components of the major histocompatibility complex (HLA-I or B2M) and of the response to IFNγ (such as JAK2) which are mutually exclusive and can affect up to one fifth of the NSCLCs. The participation of other gene alterations, such as those of common oncogenes and tumor suppressors, and of the epigenetic alterations will also be discussed, in detail. Finally, we discuss the potential use of the tumor's genetic profile to predict sensitivity to ICIs.

Abstract B42: Identification of CPT1A as a novel driver of proliferation in luminal breast cancer

Abstract Clinically, approximately two-thirds of the nearly 250,000 breast cancer cases diagnosed each year in the United States are hormone receptor-positive, luminal breast tumors. Slower-growing luminal breast tumors are often successfully treated using endocrine-based therapies, resulting in a relatively good prognosis for these patients. However, more highly proliferative luminal tumors, even with the recent approval of CDK4/6 inhibitors, are often resistant, or become resistant, to current therapies, leading to a worse outcome for these women. Therefore, it is necessary to identify genetic events responsible for tumorigenesis and to identify potential druggable genetic alterations and/or pathways in order to improve clinical outcome. To address this question, we recently developed and used an innovative genomics-based strategy to interrogate orthogonal genome-wide data from more than 2,500 patients from the TCGA and METABRIC studies. By using an 11-gene mRNA-based gene expression signature of proliferation as a conceptual framework, we identified DNA copy number alterations and somatic mutations associated with luminal breast cancer proliferation. In order to identify the subset of amplified genes that are essential for cell viability, we analyzed a dataset of 27 breast cancer cell lines with available mRNA expression and genome-wide shRNA proliferation data. By integrating DNA copy number and shRNA analyses, we identified amplified carnitine palmitoyltransferase 1A (CPT1A) as a novel driver of proliferation in luminal breast cancer. CPT1A was found to be amplified in 40.2% of highly proliferative (top quartile) luminal tumors compared to 24.7% of all other samples and CPT1A DNA copy number status correlated with mRNA expression levels. Importantly, CPT1A was shown to be essential in luminal breast cancer cell lines with a high proliferation signature score. By analyzing RPPA data from luminal breast tumors, we confirmed CPT1A amplified tumors have increased expression of protein markers of proliferation. Since CPT1A is the rate-limiting enzyme responsible for fatty acid import into the mitochondria during fatty acid β oxidation (FAO), these data indicate that highly proliferative luminal tumors may utilize FAO as a prominent energy source. Interestingly, our analyses of RNA-seq data (n=1,031) demonstrated that highly proliferative luminal tumors have significantly higher CPT1A mRNA expression than either basal-like, which are largely synonymous with TNBC, or less proliferative luminal tumors, suggesting that these tumors may have a greater dependency on CPT1A activity. CPT1A has two major variants: cytoplasmic Variant 1 is the rate-limiting enzyme responsible for fatty acid import into the mitochondria during FAO while nuclear Variant 2 is purported to mediate histone deacetylase activity. Analyses of RNAseq data (n=819) demonstrated that Variant 1 represents >99.2% of CPT1A mRNA transcripts in a given tumor, suggesting the FAO-associated role of CPT1A is the more critical function of this protein in breast cancer. Consistent with this observation, we determined that breast cancer cell lines with high CPT1A protein expression are significantly more sensitive to etomoxir, a CPT1A-specific inhibitor known to repress FAO, when compared to cell lines with low CPT1A protein levels. Finally, we determined that under glucose-depleted conditions, cell lines with high CPT1A expression showed no difference in etomoxir response whereas cells with low CPT1A become significantly more sensitive. Collectively, these data suggest that CPT1A amplification and overexpression contribute to proliferation. Given that a number of approved or experimental drugs target CPT1A or FAO, our data suggest that CPT1A and/or FAO may represent a novel therapeutic target for luminal breast cancer. Citation Format: Kristina Mastrioanni, Kimberly Parker, Michael L. Gatza. Identification of CPT1A as a novel driver of proliferation in luminal breast cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Breast Cancer Research; 2017 Oct 7-10; Hollywood, CA. Philadelphia (PA): AACR; Mol Cancer Res 2018;16(8_Suppl):Abstract nr B42.

Abstract 2601: Clinical outcomes of pts with advanced solid tumors treated according to NGS guided-therapy in a Brazilian cancer center

Abstract Introduction: Next Generation Sequencing (NGS) is a key tool since it unreveals genetic alterations (GA) with potential for targeted-therapy, frequently, not detected for conventional methods done previously. Nevertheless, avaiability of these drugs is a major concern and, additionally, oncologists are challenged to deal with such a huge amount of findings whose clinical significance and sensibility to those drugs are, many times, uncertain. Is broadly unknown, except for isolated initiatives, the correlated clinical outcomes of targeted-therapy guided by NGS. Methods: In this retrospective study, we describe clinical outcomes of 34 pts with advanced solid tumors (tu) treated in a single Cancer Center in Brazil according to therapies guided by NGS. All tests were performed using Illumina HiSeqs of Foundation Medicine (FM). The f/u was obtained from our electronic charts. Results: From apr/14 to oct/17, 34 pts were identified, 13M/21F, mean 58y/o. Histologies were: 7 NSCLC, 10 mCRC, 3 pancreatic, 3 breast and 11 varied. In 26 / 34 pts (76%), druggable GA were identified and 17 (50%) were treated with NGS-guided therapies - 14 with TKi and 3 with immunotherapy (imm). Importantly, 8/ 17 pts (47%) experienced clinical benefit (DE, PR, CR) by RECIST 1.1 (1 melanoma, 1 colon, 6 lung adeno). Seven (41%) had PD in the first control in 4-8w (mCRC treated with cobimetinib because of TP53 G12D; mCRC-trastuzumab-HER2 R678Q; breast-olaparibe-BRCA-2 T431fs*20; breast-EVE-PIK3CA E545K; endometrium-EVE-PIK3CA C604R/PIK3CA E81K and PTEN R233*/PTEN S229; mCRC-trastuzumab+lapatinib-HER2 amplification V777L; HCC-EVE-IKBKE amplification and PTEN loss exon 2-9), 1 had just started the therapy (BRCA-2 mutated pancreatic adeno with olaparib) and 1 died before restaging (HER2-mut mCRC treated with chemo+trastuzumab). The DoR was better in those with anti-ALK (3 pts, 18 mo) and anti-EGFR (1 pt, 6 mo ongoing response) and in those with TMB-I/high treated with imm (1 melanoma with TMB27 and 2 lung adeno with TMB 18 and 8). Amongst the 10 pts with evaluable TMB values, 6 were TMB-I/high and, in those, was found a high frequency of mutations: mean of 7). Curiously, 1 pt with mCRC with Kras G12V mutation treated with Trametinib, according to FM recommendations, experienced SD for 12 mo. Finally, 7 pts had samples tested by local companies using other methods (IHC, FISH, PCR) and in 5 (14% of all 34) it did not identified the GA observed by NGS. Conclusion: NGS identified a significant amount of GA otherwise not detected by conventional tools, changed management and resulted in improved clinical benefit, especially for those with lung adeno treated by anti-ALK/EGFR and those with TMB-I/high treated by imm. Addionally, this study suggests that tu with higher amount of GA in NGS are prone to harbor TMB-I/high and also confirms the low responsiveness of solid tumors to EVE even with driver-mutations in the PI3K-Akt-mTor pathway. Citation Format: Marcos Andre Costa, Marcelo Santos, Roberto Abramoff, Renata D'alpino, Carlos Teixeira, Ariel Kann, Jacques Tabacof, Riad Younes. Clinical outcomes of pts with advanced solid tumors treated according to NGS guided-therapy in a Brazilian cancer center [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2601.

Molecular Testing of Lung Cancers.

Targeted therapies guided by molecular diagnostics have become a standard treatment of lung cancer. Epidermal growth factor receptor (EGFR) mutations and anaplastic lymphoma kinase (ALK) rearrangements are currently used as the best predictive biomarkers for EGFR tyrosine kinase inhibitors and ALK inhibitors, respectively. Besides EGFR and ALK, the list of druggable genetic alterations has been growing, including ROS1 rearrangements, RET rearrangements, and MET alterations. In this situation, pathologists should carefully manage clinical samples for molecular testing and should do their best to quickly and accurately identify patients who will benefit from precision therapeutics. Here, we grouped molecular biomarkers of lung cancers into three categories—mutations, gene rearrangements, and amplifications—and propose expanded guidelines on molecular testing of lung cancers.

Abstract 75: Patient-specific genomic profiling for advanced cancers in young adults

Abstract Purpose: Large-scale cancer genome analyses have uncovered the genetic landscape of common cancers. However, rate tumor types in adolescent, young adult (AYA) have not been well-characterized genetically. We performed a pilot study of patient-specific cancer genome analysis to know whether we can analyze individual cancer genome without the existing large-scale genomics data of same tumor types in advanced and rare AYA tumors. Methods: The patients were prospectively enrolled from a medical oncology clinic in a university hospital. Tissue samples of the study patients were acquired from the fresh tissue, formalin-fixed paraffin-embedded (FFPE) tissue and cancer cells from pleural effusion as well. Cancer genome data was acquired using massive parallel sequencing technique from seven AYA patients with advanced solid tumors. WES and WTS data were generated from 6 tumors with matched normal and 4 tumors, respectively. Somatic alterations of cancer genome were classified with 2 distinct categories with heuristic ways and level 1 to 3 depends on differences of genetic alterations based on clinical and biological relevance, and mutation context. Results: Each different tumor types of 7 study patients were as follows; atypical prostate cancer (AYA01, 30 years old), olfactory neuroblastoma (AYA02, 30 years old), tongue cancer (AYA04, 33 years old), urachal carcinoma (AYA06, 32 years old), germ cell tumor (AYA07, 21 years old), lung cancer (AYA09, 34 years old) and liposarcoma (AYA10, 33 years old). Patients with GCT and tongue cancer had higher mutation rate based on WES data; 476 and 97 non-synonymous somatic nucleotide variations respectively. However, there were only 13∼16 non-synonymous somatic nucleotide variations and 7∼18 indel somatic variations in patients with other four cancers. We identified one level-1 (strong) oncogenic alterations and eight level-1 tumor suppressor alterations as well as 19 level-2 (moderate) and level-3 (modest) alterations were found from 5 WES data (we excluded 1 hypermutated germ cell tumor sample). Some level-1/2 alterations were considered as targetable by developed or developing drugs. Each tumor was characterized by tumor-unique manner. AYA01 was characterized with concurrent tumor suppressor alterations of RasGAP family genes (NF1 and RASA2), AYA02 with chromosome-level copy number alteration and CDKN2C stopgain mutation, AYA04 with concurrent oncogenic (AMER3) and tumor suppressor (FAT1, LGR6, MSX1) alterations of Wnt signaling, AYA06 with KRAS mutation and AYA09 with a fusion of EML4-ALK. Conclusion: Our study showed that we can characterize rare AYA tumors with WES or WTS data without pre-existing large-scale genomics data of same tumor types, although there are much hurdles in identifying major driver and/or druggable genetic alterations. Citation Format: Soojin Cha, Jeongeun Lee, Jong-Yeon Shin, Ji-Yeon Kim, Jong-Il Kim, Se-Hoon Lee. Patient-specific genomic profiling for advanced cancers in young adults. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 75. doi:10.1158/1538-7445.AM2015-75

Abstract LB-32: Patient-derived xenograft (PDX) paraclinical platform to guide precision medicine in urothelial cancer

Abstract BACKGROUND: The overall efficiency in personalizing medicine by pairing targeted therapy with specific genomic alterations identified through omics technologies is disappointing as current computational biology methods have difficulty in distinguishing “driver” alterations from “passenger” ones. We have developed PDXs that share essentially the same genetic background as the parental human cancer, and use this platform to screen for most effective drugs and perform mechanistic study of drug resistance in order to guide design of precision medicine in urothelial cancer. MATERIALS AND METHODS: PDXs were developed by direct implantation of fresh clinical tumor specimens in immunocompromised NSG mice (The Jackson Laboratory, West Sacramento, CA). Whole exome, transcriptome and microRNA sequencing was performed, followed by computational biological analysis to identify druggable genetic alterations that were further validated with immunofluorescence (IF) and immunohistochemical (IHC) staining. Efficacy tests to determine response to treatment by targeted therapy, together with analyses to determine the mechanisms of resistance were performed in NSG mice carrying PDXs. RESULTS: The engraftment rate of PDX was 41 % (13/42). The fidelity between the PDXs and the parental patient cancers was confirmed with cell morphology, tissue microarray IHC/IF staining and mutation profiles. Cell morphology was maintained during passaging in both the subcutaneous and orthotopic PDXs. All PDXs had multiple druggable genetic alterations. Of the first 8 PDXs, 2 harbored PIK3CA mutations (H1047R at the kinase domain and D549Y at the helical domain); 5 had ERBB2 mRNA overexpression determined by transcriptome sequencing, and protein overexpression (3+ by IHC), and 5 had EphB4 overexpression. sEphB4-HSA is an experimental drug developed by Gill (co-author). In PDXs overexpressing EphB4, blockade of EphB4 with sEphB4-HSA significantly prolonged the progression-free survival (PFS) from 9.5 days in the control group to 16.7 days (p=3.17 X 10e-5). Western blot, IHC, and deep sequencing analyses in PDX tissue specimens collected before/during treatment, and after the development of resistance were used to determine the mechanism of resistance. CONCLUSION: PDXs can potentially serve as a para-clinical model for drug efficacy screening, study of resistance mechanisms, and drug development. Citation Format: Chong-xian Pan, Hongyong Zhang, Clifford Tepper, Paramita Ghosh, Sheri Kuslak-Meyer, Susie Airhart, Parkash Gill, David Gandara, Edison Liu, Ralph de Vere White. Patient-derived xenograft (PDX) paraclinical platform to guide precision medicine in urothelial cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-32. doi:10.1158/1538-7445.AM2014-LB-32