PIK3CA Alterations Research Articles

Undetectable circulating tumor DNA levels correlate with low risk of recurrence/metastasis in postoperative pathologic stage I lung adenocarcinoma patients

ObjectivesThe application of circulating tumor DNA (ctDNA) monitoring after resection in pathologic(p) stage I lung adenocarcinoma (LUAD) patients remains controversial and it is of great clinical interest to decipher the difference of genetic features between ground-glass opacity (GGO) and solid nodules (non-GGO) subgroups. We aim to assess the utility of ctDNA in tracking early recurrence or metastasis following surgery and reveal the genetic differences between GGO and non-GGO. Materials and methodsTumor tissues and matched postoperative plasma samples were collected from a total of 82 (p)stage I LUAD patients. Comprehensive genomic profiling was performed using capture-based hybrid next generation sequencing by targeting 422 cancer relevant genes. ResultsEGFR and TP53 represent commonly mutated genes in this cohort of (p)stage I lung adenocarcinoma, followed by alterations in ALK, PIK3CA, STK11 and MYC. For a median follow-up period of 22.83 months after surgery, 65 out of 67 ctDNA-negative patients remained progression-free, while 3 out of 15 ctDNA-positive patients progressed [P = 0.040; positive predictive value = 0.20, 95 % confidence interval (CI), 0.04−0.48; negative predictive value = 0.97, 95 % CI, 0.9–1]. With time-dependent Cox regression analysis, we observed that ctDNA positivity significantly correlated with increased probability of early tumor recurrence or metastasis (P = 0.02, HR=8.5). Further comparison between GGO and non-GGO subgroups indicated the frequency of TP53 mutations in non-GGO was markedly higher than that in GGO (47 % vs 21 %, P < 0.05). Pathway analysis showed the epigenetic regulation pathway was more frequently affected in GGO subgroup, while impaired apoptosis/cell cycle pathway was more enriched in non-GGO LUADs. ConclusionsOur longitudinal ctDNA monitoring data showed that undetectable ctDNA may predict low risk of tumor recurrence or metastasis in postoperative (p)stage I LUAD patients, while it requires further investigation on how robust the positive ctDNA results could predict tumor relapse in these patients. Clinical registration numberNCT03172156.

Vulvar Squamous Cell Carcinoma: Comprehensive Genomic Profiling of HPV+ Versus HPV- Forms Reveals Distinct Sets of Potentially Actionable Molecular Targets.

PURPOSEVulvar squamous cell carcinoma (vSCC) encompasses two predominant variants: one associated with detectable high-risk strains of human papillomavirus (hrHPV) and a second form often occurring in the context of chronic dermatitis in postmenopausal women. Genomic assessment of a large-scale cohort of patients with aggressive vSCC may identify distinct mutational signatures.MATERIALS AND METHODSTumor samples from a total of 280 patients with vSCC underwent hybridization capture with analysis of up to 406 cancer-related genes. Human papillomavirus (HPV) sequences were detected by de novo assembly of nonhuman sequencing reads and aligned to the RefSeq database. Immunohistochemistry for programmed death-ligand 1 (PD-L1) was assessed.RESULTSOne hundred two of 280 vSCCs (36%) contained hrHPV sequences, predominantly HPV 16 (88%). The HPV-positive (HPV+) group was significantly younger (median age, 59 v 64 years; P = .001). Compared with HPV-negative (HPV–) vSCCs, HPV+ tumors showed more frequent pathogenic alterations in PIK3CA (31% v 16%; P = .004), PTEN (14% v 2%; P < .0001), EP300 (14% v 1%; P < .0001), STK11 (14% v 1%; P < .0001), AR (5% v 0%; P = .006), and FBXW7 (10% v 3%; P = .03). In contrast, HPV– vSCCs showed more alterations in TP53 (83% v 6%; P < .0001), TERTp (71% v 9%; P < .0001), CDKN2A (55% v 2%; P < .0001), CCND1 amplification (22% v 2%; P < .0001), FAT1 (25% v 4%; P < .0001), NOTCH1 (19% v 6%; P = .002), and EGFR amplification (11% v 0%; P < .0001), as well as a higher rate of 9p24.1 (PDL1/PDL2) amplification (5% v 1%) and PD-L1 immunohistochemistry high-positive tumor staining (33% v 9%; P = .04).CONCLUSIONComprehensive molecular profiles of vSCC vary considerably with hrHPV status and may inform patient selection into clinical trials. Sixty-one percent of HPV+ vSCCs had a pathogenic alteration in the PI3K/mTOR pathway, whereas HPV– vSCCs showed alterations in TP53, TERTp, CDKN2A, CCND1, and EGFR, and biomarkers associated with responsiveness to immunotherapy.

Abstract PR04: RNA sequencing of salivary mucoepidermoid carcinoma reveals distinct molecular profiles based on histologic grade

Abstract Introduction: The presence of the t(11;19) chromosomal translocation and its CRTC1-MAML2 (C1/M2) fusion product occurs frequently in salivary mucoepidermoid carcinoma (MEC) and has been correlated with a favorable prognosis. Recently, whole-exome sequencing studies found genomic alterations in MEC to be infrequent, which supports the role of C1/M2 as an early driver event. However, when present, TP53 and PIK3CA alterations may represent alternative driver mutations in fusion-negative cases. To better understand the MEC transcriptional landscape by fusion status and histologic grade, we performed RNA deep sequencing. Methods: 16 parotid MEC (8 low-grade and 8-high grade) tumors, 12 adjacent normal, and 6 age-matched normal parotid tissues were retrospectively analyzed from UNC Surgical Pathology specimens collected from 2000-2019. H&amp;E and mucicarmine stained slides were independently reviewed by pathology for diagnostic confirmation. The Air Force Institute of Pathology (AFIP) classification system by Ellis and Auclair was utilized for histologic grading. RNA was extracted from FFPE tissue sections and quality checked prior to library preparation. RNA-seq (HS2500) was then performed on all samples and computational methods were used to determine C1/M2 fusion status. Gene set variation analyses (GSVA) were performed on genes differentially expressed between MEC tumor versus normal, or according to tumor grade, and/or fusion status. Results: Compared to matched normals, MEC tumors display significant changes in ERBB2 and PI3K signaling cascades (FC=-3.2, p=0.049). We also noted significant upregulation of inflammatory and apoptosis markers including interferon gamma, IL-4, NOS1, and programmed cell death (p&amp;lt;0.05). Moreover, compared to low-grade, high-grade MEC tumors had significant upregulation of angiogenesis pathways, including the VEGF-VEGF receptor signaling network (FC=4, p=0.007) and activation of developmental pathways such as TGF-β signaling (FC=3.62, p=0.015). Notably, we report the expression of C1/M2 fusion transcripts in both low- and high-grade MEC. Further, p53 signaling pathway is downregulated in C1/M2 fusion-positive cases compared to fusion-negative cases (p&amp;lt;0.05). Discussion: This is the first study to perform transcriptome analyses of salivary MEC comparing to normal parotid tissue, while accounting for histologic grade. While C1/M2 fusion status is considered pathognomonic of low-grade MEC, we identified the expression of C1/M2 fusion transcripts in high-grade MEC. This confirms recent reports challenging the prognostic value of fusion status. Our study also confirms that reprogramming TP53 and PI3K signaling is a universal feature of MEC and epigenetic mechanisms may repress p53 in the absence of TP53 driver mutations seen in fusion negative tumors. Lastly, we report that VEGF and TGF-β pathway activation are associated with cancer pathogenesis in high-grade MEC. The evaluation of anticancer therapies targeting these pathways should be evaluated further. Citation Format: Siddharth Sheth, Renee Betancourt, Joel Parker, Kshitij Sharma, Adele Musicant, Wendell Yarbrough, Trevor Hackman, Antonio Amelio. RNA sequencing of salivary mucoepidermoid carcinoma reveals distinct molecular profiles based on histologic grade [abstract]. In: Proceedings of the AACR-AHNS Head and Neck Cancer Conference: Optimizing Survival and Quality of Life through Basic, Clinical, and Translational Research; 2019 Apr 29-30; Austin, TX. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(12_Suppl_2):Abstract nr PR04.

Clinical potential of ctDNA-based TMB in small cell lung cancer recieving chemoradiotherapy.

3536 Background: Small cell lung cancer (SCLC) is an aggressive tumor with poor prognosis. Chemotherapy and / or radiotherapy is the main choice of SCLC treatment. Circulating tumor DNA (ctDNA) has received substantial attention in recent years owing to the potential of patient stratification and monitoring. Here, we assessed the value of prediction and prognosis using ctDNA in SCLC. Methods: SCLC patients (pts) with limited-stage disease (LD) receiving chemoradiotherapy and extensive-stage disease (ED) receiving chemotherapy were enrolled. Baseline plasma samples were collected for NGS using a 1021-gene-panel. Mutational features and blood-based tumor mutation burden (bTMB) were analyzed using ctDNA. pyClone software was used to cluster the mutations. The mutations in the cluster with the highest cancer cell fraction (CCF) were defined as clonal mutations. Progression-free survival (PFS) was followed. Results: 58 SCLC pts (35 LD and 23 ED) and 58 plasma samples were enrolled. Smoking pts accounted for 84% (49/58). In all samples, recurrent genes were TP53 (86%), RB1 (57%), LRP1B (34%), CREBBP (26%), and MLL3 (22%). The median of bTMB and clone count were 7.9 [0-26] and 7 [0-25]. Significant higher bTMB and clone count were observed in ED pts compared with LD (Mann Whitney test, p = 0.019 and p = 0.041, respectively). Mutated CREBBP (10/23 ED versus 5/35 LD) was enriched in ED (Fisher exact test, p = 0.017 and OR = 0.223). Mutations in NOTCH signaling pathway were enriched in ED (l6/23 ED versus 13/35 LD, p = 0.031, OR = 0.265). In LD group, there were trend toward prolonged PFS in pts with higher bTMB(p = 0.065), and pts with higher clonal bTMB (cbTMB) exhibited significant longer PFS (p = 0.016, HR 0.37, 95% CI [0.12-1.11]). Patients with alteration in PIK3CA showed shorter PFS than wild type (p &lt; 0.001, HR 0.11, 95% CI [0-2.86]). There were no significant difference in median PFS in LD stage pts with any detectable pathway alterations. Whereas, LD pts whose ctDNA contained RTK-RAS signaling pathway alterations exhibited shorter PFS than pts without those alterations (p = 0.135). In ED pts, NOTCH1 gene wild type displayed longer PFS than mutant type (p = 0.036, HR 0.38, 95% CI [0.1-1.53]). There were no difference in PFS between pts with higher and lower bTMB and cbTMB. Conclusions: ctDNA can characterize the mutational feature of SCLC. There are differences in the molecular characteristics between ED and LD pts. Clonal bTMB is a potential prognostic biomarker for LD SCLC chemoradiotherapy. The prognostic marker of ED chemotherapy is different from LD.

Acquired genomic alterations in circulating tumor DNA from patients receiving abemaciclib alone or in combination with endocrine therapy.

3519 Background: An understanding of the mechanisms of acquired resistance to CDK4 &amp; 6 inhibitors, either alone or with endocrine therapy (ET), is an unmet need. Abemaciclib is a CDK4 &amp; 6 inhibitor approved for treatment of HR+, HER2- advanced breast cancer (ABC). Here we evaluated acquired genomic alterations detected in circulating tumor DNA (ctDNA) from patients (pts) treated with abemaciclib + nonsteroidal aromatase inhibitor (AI) or placebo + AI in MONARCH 3 or abemaciclib monotherapy in nextMONARCH 1. Methods: MONARCH 3 randomized postmenopausal women with HR+, HER2- ABC with no prior systemic therapy in the advanced setting to abemaciclib (150 mg Q12H) or placebo + AI. nextMONARCH 1 randomized women with HR+, HER2- metastatic breast cancer who had progressed on or after prior ET and CT to abemaciclib (150 mg Q12H) + tamoxifen, abemaciclib (150 mg Q12H), or abemaciclib (200 mg Q12H) + loperamide. Plasma from pts in the abemaciclib or placebo + AI arms (MONARCH 3) or abemaciclib monotherapy arms (nextMONARCH 1) was analyzed by the Guardant360 assay to identify potential tumor-related genomic alterations including point mutations, indels, amplifications, and fusions acquired at EOT in comparison with baseline. Results: For MONARCH 3, commonly acquired alterations at EOT included ESR1 (17%), TP53 (10%), EGFR (8%), FGFR1 (7%), and PDGFRA (7%) in the abemaciclib + AI arm, and ESR1 (31%), TP53 (10%), and BRCA1 (7%) in the placebo + AI arm. Acquired alterations more frequent for abemaciclib + AI pts included RB1 (6%), MYC (5%), and AR (5%), compared to 0% in the placebo + AI arm (p = 0.008 RB1; p = 0.015 MYC or AR). In contrast, acquired ESR1 alterations were less frequent with abemaciclib + AI vs placebo + AI (17% vs 31%, p = 0.038). In nextMONARCH 1, the most commonly acquired alterations with abemaciclib monotherapy were in TP53 (10%), EGFR (9%), RB1 (9%), MYC (9%), and MET (8%). In addition, acquired alterations in ESR1 (6%) and AR (3%) were also found. PIK3CA alterations were not frequently acquired (abemaciclib + AI 1%, placebo + AI 6%, abemaciclib monotherapy 5%). Conclusions: Acquired genomic alterations potentially associated with emerging mechanisms of resistance to abemaciclib alone or in combination with AI may include RB1, MYC, or AR alterations, while the acquisition of ESR1 alterations was less common in pts treated with abemaciclib + AI compared to placebo + AI. These findings are hypothesis-generating and provide insight into mechanisms of resistance to abemaciclib vs ET. Clinical trial information: NCT02246621, NCT02747004 .

Patho-genomic feature of PI3K / AKT / mTOR pathway mutations in Chinese patients with non-small cell lung cancer.

e21049 Background: Inhibition of PI3K/AKT/mTOR pathway has emerged as a promising anticancer strategy. Measuring genetic alterations in PI3K/AKT/mTOR pathway are critical for clinical decision making for patients with lung cancer. This study aims to analyze PI3K/AKT/mTOR pathway-related gene mutations in non-small cell lung cancer (NSCLC) in a consecutive cohort. Methods: 536 surgically resected tumor tissues were collected. Target region capture sequencing for 508 cancer-related genes was conducted on MGI-seq 2000 platform. Results: PI3K/AKT/mTOR pathway genes were mutated in 120 samples (22.4%). Female or older patients displayed higher mutation incidence of PI3K/AKT/mTOR pathway (P = 0.06 for female, P = 0.05 for patients ≥60 y). In NSCLC samples, mutations mainly occurred in NF1/2(18/499), PTEN (7/499), MTOR (4/499), TSC1/2(18/499), PIK3CA (26/499), PDK1 (2/499), AKT1/2/3 (5/499) and KRAS (49/499) genes. Genetic alterations of AKT2 (p = 0.003), mTOR (P = 0.03), PTEN (P = 0.005) and PIK3CA (P &lt; 0.001) are preferred to occur in squamous NSCLC. PIK3CA variants (p.E542K, p.E545K, p.Q546K) were more enriched in non-squamous NSCLC, while amplification of PIK3CA was more prevalent in squamous NSCLC (P = 0.038). Histologically, TSC1/2 mutations were more correlate with micro-invasion subtype in non-squamous NSCLC (P = 0.002). Meanwhile, PIK3CA alterations, especially, amplification occurred more often in keratinizing squamous NSCLC (P &lt; 0.001). In addition, four patients with biallelic NF1 loss of function mutations were exclusively found in non-squamous NSCLC without co-occurrence of mutation in known driver genes like EGFR or TP53, which indicated biallelic loss of function of NF1 might be sufficient to drive oncogenesis of non-squamous NSCLC. Conclusions: PI3K/AKT/mTOR pathway alterations present a heterogenous distribution across various types of lung malignancies especially in NSCLC. As more small molecule inhibitors developed to target PI3K/AKT/mTOR pathway, this study might shed light on further clinical investigation of these lately developed therapeutic approaches. [Table: see text]

Patient-matched tissue and liquid biopsies identify shared and acquired genomic alterations in breast cancer.

1050 Background: Liquid biopsy-based comprehensive genomic profiling (CGP) is a minimally invasive approach to potentially identify truncal driver alterations and mechanisms of acquired resistance. However, there are limited data comparing solid tissue and plasma biopsies from the same patient in breast cancer. Methods: CGP was performed on matched tumor tissue and plasma samples from 444 patients with breast cancer (Foundation Medicine, Inc.) to identify short variant mutations and rearrangements in at least 62 genes. ER/PR/HER2 status was abstracted for a subset of patients (n = 273). Patients were primarily ER+/HER2- (58%) with fewer HER2+ (16%) and TNBC (25%). Results: Samples were temporally heterogeneous with a median time between matched tissue-plasma collection of 276 days (interquartile range 45 - 650). Positive percent agreement (PPA) to tissue biopsy was 81.5% with higher PPA for collection interval ≤1y v &gt; 1y (82.5% v 76.5%). PPA was highest in known truncal driver genes ( AKT1, PTEN, BRCA2, BRCA1, TP53, PIK3CA: 85-89% PPA). PPA of tissue to liquid biopsy was 50.5% with strong time dependence (62.4% ≤1y, 37.9% &gt; 1y), presumably due to intervening therapies. Acquired alterations in liquid biopsy were primarily known resistance mechanisms ( ESR1, NF1, ERBB2, PIK3CA, PTEN, TP53, BRCA1, and BRCA2), were often polyclonal, associated with longer collection intervals, and exhibited higher prevalence in the ER+/HER2- subtype (62% ER+/HER2-, 33% HER2+, 42% TNBC). Acquired alterations significantly enriched in ER+ cases included ESR1, NF1, ERBB2 and TP53. For PIK3CA, a PPA of 84.6% was observed with solid as baseline and 79.1% with liquid as baseline; prevalence was similar (36.5% solid, 35.6% liquid). Alpelisib companion diagnostic alterations (CDx) had a similar PPA (84.0% solid, 83.3% liquid). Overall percent agreement (OPA) for PIK3CA CDx in paired samples was 91.4% (406/444). When acquired PIK3CA alterations are observed they tend to be subclonal, co-occur with shared PIK3CA alterations, and consist of rare alterations (E726K, E453K, M1043I). Conclusions: While high PPA to tissue was observed for alterations in truncal driver genes, acquired alterations observed in plasma were frequently polyclonal, often identify potential mechanisms of therapeutic resistance and are, in part, a function of time and intervening treatments relative to tissue CGP. These results demonstrate high PPA to tissue suggesting that liquid biopsy has clinical validity in identifying truncal alterations and can also identify acquired alterations in breast cancer.

CtDNA resistance landscape of lazertinib, a third-generation EGFR tyrosine kinase inhibitor (TKI).

9601 Background: While EGFR mutant ( EGFRm) non-small cell lung cancer (NSCLC) patients usually experience improved clinical benefit with EGFR TKIs, most eventually progress. Understanding mechanisms of resistance (MoR) may allow for more personalized treatment. Lazertinib is an irreversible third generation EGFR TKI for which MoR are unknown. Obtaining sufficient tumor tissue for genotyping at progression is often difficult. Therefore, we utilized plasma ctDNA from patients treated with lazertinib to explore MoR. Methods: Plasma samples from 47 NSCLC patients in the phase 2 trial of lazertinib (NCT03046992) were collected at screening and progressive disease (PD) and underwent ctDNA NGS of 74 genes using Guarant360. All patients were positive for an EGFR Ex19del or L858R ( EGFRm) and T790M by tissue testing at screening. Acquired, nonsynonymous, characterized mutations detected in a PD sample but not in the screening sample from the respective patient were considered putative MoR, excluding aneuploidy. Patients with detectable plasma EGFRm and/or T790M at screening were evaluable. Results: ctDNA was detected in 47 (100%) screening samples and 43/45 (96%) PD samples (two failed sequencing). An EGFRm was detected in 85% of patients at screening (n = 40), 38 of which had PD ctDNA results and were included in analysis. T790M was detected in 30 patients at screening and subsequently not detected at PD in 21 of these patients, 55% of all 38 included patients. Among the ten patients with T790M detected at PD, on-target MoR were detected in 7 (18% of all included patients) including EGFR C797S (n = 3, 8%), EGFR amplification (n = 3, 8%), and EGFR T854A (n = 1, 3%). All C797S were in cis with T790M. No on-target MoR were detected in patients without T790M detected at PD. Off-target MoR were seen in 34% of patients (13/38) including mutations in PIK3CA (13%; 2 E545K, 2 E542K, 1 E81K), ERBB2 (5%; 1 D769H, 1 V777L), KRAS (3%; 1 G12C), and BRAF (3%; 1 G469A). Gene amplifications were detected in CCND1 (n = 1, 3%) , CCNE1 (n = 2, 5%) , ERBB2 (n = 1, 3%) , FGFR1 (n = 1, 3%) , MET (n = 4, 11%) , and PIK3CA (n = 1, 3%), with some patients having multiple MoR. Conclusions: The spectrum of MoR identified in this cohort of patients treated with lazertinib is similar to that reported in other third generation EGFR TKIs, but with some differences in frequencies. The most common resistance mechanisms are T790M loss and PIK3CA alterations which may address the mechanism of action. Our findings suggest putative MoR of lazertinib and show that ctDNA NGS is an effective way to identify MoR in patients progressing on targeted therapy. Clinical trial information: NCT03046992 .

Liquid biopsy: A community-based oncology practice experience.

3527 Background: Liquid biopsy is a promising and minimally invasive genetic test examining plasma circulating tumor DNA. Coupled with the rapidly developing next-generation sequencing (NGS) technologies, it holds the potential for implementation in selecting signal-matched therapeutic options. Methods: A retrospective chart review was conducted on adult patients with advanced solid tumors whose tumors were tested with Guardant360 assay, between December 2018 and December 2019. A total of 178 patients were referred for testing by 12 oncologists within a single community cancer center. Results: Referral rates varied widely (2.25% - 22%). The majority of patients (98%) were tested upfront for molecular marker evaluation, in either newly diagnosed advanced cancer patients, or in recurrent patients without enough tissue for testing. Other patients (2%) were evaluated after failure of 1 st line therapy to assess for acquired mutations. A total of 18 histological types were tested, with lung (LCa; n = 90; 50.56%), breast (BCa; n = 31; 17.42%), and colorectal (CRCa; n = 14; 7.87%) cancers being the most common types. In 86.11% of all tests (n = 180), ≥ 1 alteration was detected, while 13.89 % of tests did not reveal any tumor-related mutation, and were considered negative. The average number of alterations per test was 3.1 (±2.14; n = 481), and varied across types: CRCa (4.36), prostate cancer (2.73), BCa (2.97), and LCa (2.59), had the highest average number of alterations per test. Similarly, LCa (48.44%), BCa (19.13%), and CRCa (12.68%), harbored most of the detected somatic alterations (n = 481). Of all the alterations of practical significance (n = 457), TP53 (32.17%), PIK3CA (8.53%), EGFR (7.66%) and KRAS (7.22%), were the most commonly altered genes. Only 1 patient had a positive MSI-H status, amenable to immune-therapy. Of those with positive test results (n = 155), 31 (20%) had ≥ 1 FDA approved, target-matched therapeutic opportunity. Similarly, 71 patients (45.81%) had ≥ 1 target-matched therapeutic opportunity, outside current indications. Lastly, when no FDA-approved target-matched therapy was available (n = 39), results from liquid biopsy testing offered signal-based clinical trial opportunity in 39/39 patients. Conclusions: Implementation of NGS-based liquid biopsy testing is feasible within a community practice. In the era of precision oncology, such assays have the potential to expedite the efforts towards target-matched therapies and signal-based clinical trial opportunities. Further studies are warranted to identify the most-cost effective testing strategies.

Gastric-type adenocarcinoma of the cervix: Genomic drivers and clinical outcomes.

6030 Background: Cervical Gastric-type adenocarcinoma (CGA) is a non-HPV-associated adenocarcinoma, comprising 10% of all cervical adenocarcinomas (Park et al, 2019). The optimal management approach is unclear, given that most data in advanced cervical cancer is driven by HPV-positive disease. We summarize our experience with this rare tumor type at a large cancer center. Methods: A retrospective review was performed for all women diagnosed with CGA 6/1/2002- 7/1/2019. Patients who did not follow up after a single visit were excluded. Kaplan-Meier survival analysis was performed to determine progression-free survival (PFS) and overall survival (OS) from date of diagnosis. Tumors from a subset of patients were subjected to MSK-IMPACT targeted sequencing and analysis (Zehir et al, 2017). Results: A total of 68 women were identified; 47 met inclusion criteria. The median age at diagnosis was 52 years (range 27-83). The majority of patients were white (70%), an additional 19% were Asian. The majority of patients (60%, n=28) presented with advanced disease (FIGO 2018, stage II-IV), while 40% (n=19) were Stage I. Of note, 26% (n=12) had positive pelvic lymph nodes and 13% (n=6) had ovarian metastases at time of surgical resection. For upfront treatment: 13% (n=6) had surgery alone of whom 83% had stage 1 disease, 36% (n=17) had surgery followed by adjuvant therapy, 30% (n =14) received definitive chemo-radiation (CRT). All patients with stage IV disease 15% (n=7) received chemotherapy alone. At completion of primary treatment, 19% (n=9) of patients had persistent disease. In patients who received CCRT, 65% (n=22) recurred, the majority (64%) within 12 months of completion of upfront therapy. Pelvic recurrence was the most common site (n=14, 64%). With a median follow up time of 30 months (range 1-159), the median PFS for Stage I was 34.4 months, compared to 17.5 months in patients with Stage II-IV disease (p= 0.29). Of the 24 patients that had MSK-IMPACT, the most common mutation was TP53 (n=16, 64%) followed by mutations in the RAS pathway (n=8, 33%), PIK3CA (n=3, 12.5%), STK11 (n=3, 12.5%), and ERBB2 alterations (n=2, 8.3 %). 2 (8.3%) women enrolled on a clinical trial based on their NGS results, one targeting ERBB2 and one targeting PIK3CA. Conclusions: Consistent with prior published literature, CGA is an aggressive form of cervical cancer with poor median OS in the advanced setting. With universal HPV vaccination, HPV negative cervical cancer will represent a larger percentage of newly diagnosed cancers and further research is needed to identify the optimal management approach.

The landscape of muti-PIK3CA variations of 10,000 Chinese solid-tumor patients.

e15670 Background: PI3K/mTOR signaling pathway plays a central role in cancer biology. Our study aimed to explore the characteristics of multiple PIK3CA variations in the Chinese patient with solid tumors. Methods: Formalin fixed, paraffin embedded (FFPE) tumor samples and matched peripheral blood from 10,194 Chinese solid tumor patients were collected for next-generation sequencing based 450 genes panel assay. The testing was carried out by OrigiMed, a College of American Pathologists (CAP) accredited and Clinical Laboratory Improvement Amendments (CLIA) certified laboratory, Shanghai, China. Genomic alterations including SNV, short and long Indel, CNV, Fusion/Rearrangement were assessed. Microsatellite stable (MSS) status and TMB (tumor mutational burden) were also acquired by an NGS algorithm. Results: A total of 672 (6%) patients were identified harboring PIK3CA mutations, 93 (13.8%) of them had multiple PIK3CA mutations. Patients with multiple PIK3CA mutations included 51 females and 42 males with a mean age of 56 years old. Tumors with multiple PIK3CA mutations were composed of stage I (N = 14), II (N = 22), III (N = 18), IV (N = 25) and 14 without staging information. Cervical, breast, gastric and colorectal cancers were top ranked multiple-PIK3CA-mutant tumors (21%, 17%, 15%, 12% and 6%, respectively). TMB-H (≥10 mut/Mb) was significantly higher in multiple-PIK3CA-mutant tumors than that in single-PIK3CA-mutant tumors (60% vs 32%, P &lt; 0.001). The MSS status was similar between multiple- and single-PIK3CA-mutant tumors (14% vs 11%). The most common PIK3CA alterations in both multiple- and single-PIK3CA-mutant tumors were E545 (36% vs 28%), H1047 (19% vs 24%) and E542 (16% vs 13%). Noticeably, the frequency of R88 (16% vs 3%, P &lt; 0.0001), E726 (10% vs 0.8%, P &lt; 0.0001), E453 (10% vs 1%, P &lt; 0.0001), and M1043 (9% vs 0.8%, P &lt; 0.0001) mutations were significantly higher in multiple-PIK3CA-mutant tumors than that in single-PIK3CA-mutant tumors. The top 3 most frequent PIK3CA dual mutation combinations were E545K+E726K, H1047R+E726K and E545K+R88Q. Among the co-mutated genes, copy numbers of ERBB2 were less altered in multiple-PIK3CA-mutant tumors than single-PIK3CA-mutant tumors (1% vs 6%, p &lt; 0.05). Conclusions: This study, for the first time, revealed that approximately 13% PIK3CA positive Chinese solid tumor pts harboring multiple PIK3CA mutations. Several novel dual-PIK3CA-mutations were detected and multiple-PIK3CA-mutant pts more likely had high TMB, which may guide potential immunotherapy or PI3K-targeted therapy for multiple PIK3CA Chinese solid tumor patients.

Biomarker analysis of the phase III IPATential150 trial of first-line ipatasertib (Ipat) plus abiraterone (Abi) in metastatic castration-resistant prostate cancer (mCRPC).

182 Background: IPATential150 is a randomized trial comparing Ipat + Abi vs placebo + Abi in patients (pts) with 1L mCRPC (NCT03072238). We evaluated the potential associations between PTEN loss, genomic alterations, and country of enrollment (East Asian [EAS] vs non-EAS). Methods: Before randomization, tumor samples (archival &gt; 90%) were centrally tested for PTEN loss by VENTANA PTEN (SP218) immunohistochemistry (IHC) assay (N = 1101). Tumor genetic alterations were profiled by Foundation Medicine FoundationOne CDx (F1CDx) NGS assay (n = 735). Results: Of 1101 pts enrolled, 521 (47%) had PTEN loss by IHC. An overall 76% agreement was observed between PTEN IHC and F1CDx. TMPRSS2 fusion, alterations in TP53 and PTEN were most frequent (28%-32%). Alterations in PIK3CA, SPOP, APC, MYC, RAD21, and genes involved in DNA damage repair, including BRCA2, CDK12, ATM, had a 5%-8% prevalence. PTEN loss (IHC) tended to co-occur with genetic alterations in TP53 and TMPRSS2. The prevalence of PTEN loss in EAS vs non-EAS pts was 35% vs 50% by IHC and 15% vs 31% by F1CDx. Alterations in CDK12, BRCA2, SPOP, and MYC were more prevalent in EAS pts, whereas alterations in TMPRSS2, PTEN, and TP53 were more prevalent in non-EAS pts. Conclusions: This analysis reveals molecular heterogeneity in prostate cancer and association between potentially actionable targets. It also suggests that EAS pts with mCRPC have a genetic profile that may be at least quantitatively different from non-EAS pts. Clinical trial information: NCT03072238. [Table: see text]

Abstract P4-10-04: Clinical outcomes of alpelisib in hormone receptor-positive, human epidermal growth factor receptor-2-negative advanced breast cancer by next-generation sequencing-detected PIK3CA alteration status and phosphatase and tensin homolog loss: Biomarker analysis from the SOLAR-1 study

Introduction: Hyperactivation of the phosphoinositide-3-kinase (PI3K) pathway is a frequent event in human cancers and can be a result of phosphatase and tensin homolog (PTEN) loss, activation of receptor tyrosine kinases, or alterations in PI3K isoforms. About 40% of hormone receptor-positive (HR+), human epidermal growth factor receptor-2-negative (HER2-) breast cancers harbor alterations in the PIK3CA gene, which encodes the alpha subunit of PI3K (p110α). Mutations in PIK3CA can cause PI3K pathway hyperactivation, which may contribute to endocrine resistance. Alpelisib (ALP) is an α-selective PI3K inhibitor that, in combination with fulvestrant (FUL), demonstrated improved median progression-free survival (PFS) vs placebo (PBO) + FUL (11.0 vs 5.7 mo, respectively; hazard ratio [HR] 0.65; 95% confidence interval [CI], 0.50-0.85; P Methods: SOLAR-1 is a phase 3, randomized, double-blind study of ALP 300 mg (or PBO) once daily + FUL 500 mg every 28 days + Cycle 1 Day 15 in men and postmenopausal women with HR+, HER2- ABC. Retrospectively, PIK3CA alterations (mutations and amplifications) were assessed by NGS and PTEN levels were assessed by immunohistochemistry. PTEN loss was defined based on an H-score Results: Of 572 pts in SOLAR-1, 341 (60%) had a PIK3CA mutation in tumor tissue by PCR, valid NGS results were available for 404 (71%), and PTEN levels were available for 401 (70%). Of the pts with available PTEN results, 34 (8%) had PTEN loss. PFS results and HRs, by PIK3CA alteration status as detected by NGS and PTEN status, are shown in the Table. In pts whose tumors had any PIK3CA alteration (n=239), which included a small group of pts with alterations not detectable by the PCR-based test used for screening in SOLAR-1 (n=31), mPFS was improved in the ALP + FUL vs PBO + FUL arms. In the small group of pts (n=19) whose tumors had PTEN loss but no PIK3CA alteration, ALP showed an improved mPFS and favorable HR vs the placebo arm. In pts whose tumors had multiple alterations (n=44), HR and mPFS benefits of ALP were observed, albeit with wide 95% CIs. Conclusions: These data demonstrate the efficacy of ALP in pts whose tumors harbor PIK3CA alterations as detected by NGS, including alterations that were not detectable by the PCR-based test used in SOLAR-1. Additional analyses suggest that ALP is also effective in pts with single and multiple PIK3CA mutations and in those whose tumors do not harbor a PIK3CA alteration but do have PTEN loss; however, low patient numbers in the PTEN loss subgroups may limit conclusions. Citation Format: Dejan Juric, Fabrice Andre, Christian F. Singer, Joohyuk Sohn, Mario Campone, Sibylle Loibl, Pierfranco Conte, Hiroji Iwata, Eva Ciruelos, Ingrid A. Mayer, Albert Reising, Chong Ma, Michelle Miller, Naveen Babbar, Hope S. Rugo. Clinical outcomes of alpelisib in hormone receptor-positive, human epidermal growth factor receptor-2-negative advanced breast cancer by next-generation sequencing-detected PIK3CA alteration status and phosphatase and tensin homolog loss: Biomarker analysis from the SOLAR-1 study [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-10-04.

Abstract P2-11-07: Comprehensive tumor sequencing to identify biomarkers of palbociclib response: First report of the PROMISE study

Abstract Background: The combination of cyclin dependent 4/6 kinase inhibitors (CDK4/6i) with endocrine therapy (ET) has resulted in clinically significant improvements in progression-free survival (PFS) and overall survival (OS) in hormone-receptor (HR)-positive metastatic breast cancer (MBC). However, most patients’ disease ultimately progresses on CDK4/6i and ET. Therefore, further research is necessary to understand the mechanisms driving primary and secondary resistance. PROMISE is a multicenter prospective cohort study enrolling women with HR-positive MBC commencing treatment with palbociclib + letrozole (1st line) or palbociclib + fulvestrant (2nd Line). The study provides a comprehensive “omic” assessment of blood, tumor, urine and the fecal microbiome to identify molecular or cellular features associated with primary endocrine resistance (e.g. disease progression ≤ 12 months on treatment) and acquired resistance to CDK 4/6i. Additionally, patient derived xenografts and organoids are created to test new drug strategies designed to overcome resistance to CDK 4/6i and ET. Here, we present initial sequencing results from pretreatment biospecimens collected from PROMISE study participants. Methods: On-study tumor biopsies and blood samples were collected for DNA/RNA sequencing (TempusTM). The analyzed biospecimens were all obtained prior to initiation of palbociclib and ET. We correlated patient clinical characteristics (phenotypes) with molecular data and responses to protocol treatment. The data were analyzed using a series of cutting-edge bioinformatics pipelines for somatic and germline mutations in addition to copy number alterations (CNAs). The study database was locked for analysis on 06/20/2019. Results: We analyzed the somatic single nucleotide variants/INDELs (sSNV/INDEL) profiles across the tumor samples to determine the genes that were least likely to occur as a result of background mutation processes. Twenty-six patients had somatic copy number alterations (sCNA) and/or sSNV/INDEL in at least one of 18 genes with the most significant sSNV/INDEL profiles (p &amp;lt; 0.03) which included clinically and biologically relevant genes. The genes with the most statistically significant sSNV/INDEL mutation profiles were GATA3, PIK3CA, CDH1, and ESR1 (p &amp;lt; 0.0009). We observed a high percentage of tumors with somatic alterations in GATA3 (23% sSNV/INDEL, 15% sCNA), PIK3CA (38%, 12%), CDH1 (19%, 50%) and ESR1 (19%, 58%). ESR1 mutations were more frequent in patients receiving 2nd line treatment. Other frequently altered genes included TP53 (15%, 46%), MAP2K4 (8%, 50%), DNAAF1 (8%, 50%), and CDKN1B (8%, 35%). Further, ZNF317 and F3 were altered in 9 and 7 patients, respectively. Twenty-four samples had alterations in at least one of the CDK4/6 pathway genes (RB1, CCNE2, CCND1, CDK6, ESR1, CDKN2A, CCND3, CDK4, CDK2 and CCNE1). Four patients progressed on therapy; three of the four patients had mutations in PIK3CA, and one had a mutation in ESR1. Results of the RNA sequencing data (N=26) will be presented at the SABCS meeting. Conclusions: This is the first report of a prospective study designed to characterize the genomic landscape of ER+/HER2- MBC prior to palbociclib treatment. We observed high frequencies of known targetable alterations in PIK3CA and ESR1, including in patients that progress, which is consistent with previous reports. RNA sequencing data will be presented at the meeting. Citation Format: Ciara C O Sullivan, Krishna R Kalari, Vera J Suman, Peter T Vedell, Ann Moyer, Erin Carlson, Jason Sinnwell, Tejaswi Alaparthi, Xiaojia Tang, Kevin Thompson, Jaeyun Sung, Alvaro Moreno-Aspitia, Donald Northfelt, Minetta C Liu, Tufia C Haddad, Prema Peethambaram, Saranya Chumsri, Kathryn J Ruddy, Karthik V Giridhar, Roberto A Leon-Ferre, Paula Gill, Mohammad Ranginwala, Asad Javed, Sameer Batoo, Brendan P. McMenomy, Richard Weinshilboum, Liewei Wang, Matthew P Goetz. Comprehensive tumor sequencing to identify biomarkers of palbociclib response: First report of the PROMISE study [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P2-11-07.

Abstract P4-09-11: PIK3CA somatic alterations in 412 chinese invasive breast cancers: Higher frequency of mutant H1047R detected by next generation sequencing compared to breast cancer in caucasians

Abstract Purpose: Somatic alterations of PIK3CA gene is an important therapeutic target in breast cancer (BC). The PI3Kα-specific inhibitor alpelisib was remarkably active against PIK3CA-mutated, HR-positive/HER2-negative BC in the SOLAR-1 trial. We hypothesized that PIK3CA alterations in Chinese BC patients across different molecular subtypes might differ from other ethnic groups and this information would be useful for selecting anti-PI3K therapy. Methods: The molecular profile of the PIK3CA gene was analyzed in 412 Chinese untreated invasive BC patients with ER/HER2 molecular subtypes, using a 540 gene next-generation sequencing panel. The results were compared to the molecular profile of Caucasian breast cancer patients in The Cancer Genome Atlas(TCGA-white). Results: Compared to wild type, PIK3CA alterations were more frequent in the ER+ subtype (49.3%, p=0.024), and tumors with low ki67 proliferation (58.3%, p=0.007) and low histological grade (grade I/II/III 80%, 53.4%, 35.9%, p&amp;lt;0.001). Compared to TCGA Caucasian race (TCGA-white), Chinese BC patients had a higher alteration frequency (45.6% vs. 34.7%, p&amp;lt;0.001). The p.H1047R mutation predominantly occurred among three hot spots (p.E545K, p.E542K and p.H1047R) for Chinese BC compared to that of the TCGA-white cohort (66.1% vs 43.7%, p=0.01). Nine novel mutation sites of PIK3CA were observed in the Chinese cohort that was absent among Caucasian BC patients. Among the four ER/HER2 molecular subtypes, ER+/HER2+ tumors harbored the most PIK3CA alterations (51.6%), while ER-/HER2- harbored the least (30.0%). However the latter presented the highest frequency of copy number amplification (19.05%). Conclusion: PIK3CA alterations prevail in nearly half of Chinese BC patients and has some different molecular features compared to that of Caucasian BC patients. The PIK3CA distribution patterns differed among four ER/HER2 subtypes. The results provide more insights for evaluating the results of PIK3CA inhibitors. Citation Format: Minghan Jia, Ning Liao, Bo Chen, Guochun Zhang, Yulei Wang, Xiaoqing Chen, Liping Guo, Li Cao, Hsiaopei Mok, Chongyang Ren, Kai Li, Cheukfai Li, Lingzhu Wen, Jiali Lin, Guangnan Wei, Zhou Zhang, Charles M Balch. PIK3CA somatic alterations in 412 chinese invasive breast cancers: Higher frequency of mutant H1047R detected by next generation sequencing compared to breast cancer in caucasians [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-09-11.

Abstract OT2-06-01: A phase-2 trial of neoadjuvant alpelisib and nab-paclitaxel in anthracycline refractory triple negative breast cancers with PIK3CA or PTEN alterations

Abstract Background:While TNBC patients with pathologic complete response (pCR) or residual cancer burden (RCB-I) have good survival rates, those with extensive residual disease (RCB-II or RCB-III) after neoadjuvant chemotherapy (NACT) have poor prognosis. Patients without response to their first chemotherapeutic regimen have very low chance (5%) of achieving pCR. We have created a biomarker-driven drug development strategy, ARTEMIS (A Randomized, TNBC Enrolling trial to confirm Molecular profiling Improves Survival) to identify novel targeted therapies focusing on chemo-insensitive disease. The PI3K/Akt/mTOR pathway is a critical regulator of cell growth, apoptosis, metabolism in cancer cells and its activation is known to play an important role in resistance to chemotherapy. Clinical studies have shown that the combination of PI3K pathway inhibitors with taxanes are associated with improvement in response rates and survival. Alpelisib is approved for hormone receptor positive, HER2 negative, metastatic breast cancer patients with activating PIK3CA mutations. This study hypothesizes combination of alpelisib with nab-paclitaxel will improve PCR/RCB-1 in treatment-resistant early stage TNBC patients with activating PIK3CA or PTEN loss of function alterations. Trial Design: This will be a non-randomized, two- arm, open label, phase II study to evaluate the clinical activity of alpelisib and nab-paclitaxel in high-risk TNBC patients with PIK3CA or PTEN alterations with refractory or suboptimal response to anthracycline-based NACT. Eligibility criteria Stage 1-3 TNBC defined as ER&amp;lt;10%; PR&amp;lt;10% and HER2 negativeActivating alterations in PIK3CA, or PTEN lossResidual primary tumor size of at least 1.0 cm or evidence of lymph node involvement by imaging (ultrasound or MRI) after anthracycline-based NACTECOG performance status ≤1Normal organ and marrow functionReceived at least one dose of an anthracycline-based NACT (eligible if therapy was discontinued due to disease progression or therapy intolerance) Specific aims To determine if alpelisib in combination with nab-paclitaxel will improve rates of pathologic response (pCR/RCB-0 or RCB-I) from 5% to 20% in patients with chemotherapy insensitive TNBC with activating PIK3CA alterations (Arm-1) or PTEN loss of function alterations (Arm-2)To determine the radiographic response rate for alpelisib in combination with nab-paclitaxel in chemotherapy insensitive TNBC with PIK3CA (Arm-1) or PTEN (Arm-2) alterationsDetermine toxicity of alpelisib in combination with nab-paclitaxel given in the neoadjuvant settingDetermine progression free survival (PFS) at 3 years for patients treated with alpelisib in combination with nab-paclitaxel given in the neoadjuvant setting Statistical methods Primary objective is to determine if alpelisib in combination with nab-paclitaxel in the neoadjuvant setting will improve rates of pathologic response (counting pCR/RCB-0 or RCB-I as response) from 5% (control) to 20% (experimental arms). A Simon two-stage design will be employed with 12 patients in the first stage. If at least one patient has pCR/RCB-1, 25 more patients will be added for a total of 37 patients (alpha = beta = 10%). This design has a 54% probability of early termination after the first stage if the true pCR or RCB-I probability is 5%. Contact information for people with a specific interest in the trial sdamodaran@mdanderson.org Citation Format: Senthil Damodaran, Jennifer K Litton, Kenneth R. Hess, Colleen T. Eppig, Krzysztof J. Grzegorzewski, Funda Meric-Bernstam, Ignacio I. Wistuba, Jason B White, Gaiane M. Rauch, Rosalind P. Candelaria, Beatriz E. Adrada, Bora Lim, Stacy L. Moulder, Debu Tripathy. A phase-2 trial of neoadjuvant alpelisib and nab-paclitaxel in anthracycline refractory triple negative breast cancers with PIK3CA or PTEN alterations [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr OT2-06-01.

Abstract P4-05-04: Advanced breast cancer (ABC) in young women (YW): A comprehensive genomic profiling (CGP) study

Abstract Background: ABC in YW is an aggressive disease with a higher mortality rate than that seen in older women (OW). Pre-menopausal women face challenges in balancing treatment with reproductive health and are often excluded from promising clinical trials. Studies from NCI SEER suggest that the rate of YW breast cancer has nearly doubled in the last 40 years from 1.5 to 2.9 cases per 100,000. In this study, we compare the genomic alterations (GA) in ABC in YW vs OW across ancestry groups. Methods: Hybrid-capture based CGP of at least 315 cancer-related genes was performed on 21,440 clinically advanced breast cancers in 4,303 YW (&amp;lt;45) and 17,137 OW (≥45). Predominant ancestry was assessed using a SNP-based approach (Connelly et al, 2018). Subgroups were analyzed on histological (Invasive Ductal Carcinoma (IDC), Invasive Lobular Carcinoma (ILC) and molecular subtypes (ER-positive (ER+), HER2-amplified (HER2+), TNBC). Results: Our cohort of young women (YW) exhibit different frequencies of genetic driver events compared to older women (OW) (25 genes with a corrected p &amp;lt; 0.01) including a lower frequency of alterations in PIK3CA (23% in YW v 36% in OW, p = 8E-63), TBX3 (1.1% v 3.9%, p = 1E-16), and RUNX1 (1.2% v 2.4%, p = 6E-08) and a higher frequency of BRCA1 (8.5% in YW vs 3.2% in OW; p = 1E-32), BRCA2 (6.3% v 4.3%, p = 9E-08), and GATA3 (12.5% v 8.4%, p = 5E-16). When accounting for histological and molecular subtype, many of the same trends hold (Table 1). For example, PIK3CA alterations were observed at a significantly reduced rate in YW in ER+, HER2+, and TNBC disease. Although BRCA1 mutations are generally associated with TNBC, we observed a significantly higher frequency of alterations in YW with ILC (4.7% v 0.8%, p = 0.01) and ER+ disease (5.9% v 2.2%; p = 0.003). ESR1 point mutations were lower in YW v OW (6% v 13%, p = 3E-38), possibly reflecting differences in therapy. Unexpectedly, the distribution of ESR1 LBD alterations differed between the cohorts, with Y537S as the most common alteration in YW (40% v 24% in OW; p = 1E-06). YW with ILC showed a significantly higher rate of biopsy from the female reproductive tract (40% v 7.5% in OW, p =0.0001). TNBC tumors exhibited a higher rate of brain metastasis rate in YW (23% v 6% in OW, p = 0.003). HER2 tumors exhibited a lower rate of metastases to the skin in YW (8% v 14%, p = 0.02). No significant differences were observed in sites of distant metastasis in the ER+ subtype (all p &amp;gt;0.08). YW ABC was depleted in European ancestry (OR = 0.56, p = 2e-56) and strongly enriched in Asian and American ancestry (OR = 1.7 and 1.9, respectively, p &amp;lt; 6E-15) relative to the cohort of OW. No genes exhibited significant differences in frequency across ancestry groups (p &amp;gt;0.05). Across ancestry groups, BRCA1/2, ESR1, GATA3, and TBX3 show similar patterns between YW and OW. Patterns of metastatic tissue tropism were also largely similar across ancestry groups. significance YW v OW in groupGenefreq YWfreq OWoverallILCIDCER+HER2+TNBCPIK3CA23.1%36.3%P&amp;lt;0.0001nsP&amp;lt;0.0001P&amp;lt;0.0001P&amp;lt;0.001P&amp;lt;0.01TBX31.1%3.9%P&amp;lt;0.0001p&amp;lt;0.05P&amp;lt;0.0001P&amp;lt;0.01P&amp;lt;0.01nsRUNX11.2%2.4%P&amp;lt;0.0001nsP&amp;lt;0.01nsP&amp;lt;0.001nsBRCA18.5%3.2%P&amp;lt;0.0001P&amp;lt;0.01P&amp;lt;0.0001P&amp;lt;0.01nsP&amp;lt;0.01BRCA26.3%4.3%P&amp;lt;0.0001p&amp;lt;0.05nsnsnsnsGATA312.5%8.4%P&amp;lt;0.0001nsP&amp;lt;0.0001P&amp;lt;0.0001nsnsESR16.0%12.6%P&amp;lt;0.0001P&amp;lt;0.01P&amp;lt;0.0001P&amp;lt;0.0001P&amp;lt;0.01ns Conclusions: CGP of ABC in YW vs OW revealed consistent genetic differences across molecular and histological subtypes, which may impact targeted therapies. YW have a lower frequency of alterations in PIK3CA across all molecular subtypes, potentially limiting the utility of PI3K-alpha inhibitors, such as alpelisib. The high frequency of BRCA1/2 alterations across subtypes may predict sensitivity to PARP inhibitors. While there are differences in the relative prevalence of ancestry groups in our population, we did not observe significant differences in driver alterations or tissue tropism across these groups. Citation Format: Ethan S Sokol. Advanced breast cancer (ABC) in young women (YW): A comprehensive genomic profiling (CGP) study [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-05-04.

Abstract P4-06-07: PIK3CA-mutant breast phyllodes tumors show a uniformly aggressive histology and significant mutual exclusivity with MED12 mutation

Abstract Introduction: De-regulation of the class I phosphoinositide 3’-kinase (PI3K) pathway has long been known to contribute to the development and progression of many tumors. However, the FDA only recently approved the first selective inhibitor of PIK3CA, the p110-alpha catalytic subunit of PI3K, specifically for use in treatment of a subset of PIK3CA-mutant breast carcinomas. Given the emergence of this therapeutic class of agents, we sought to identify other subsets of breast tumors that may be driven largely by PIK3CA mutations and which therefore could be candidates for these therapies. In breast fibroepithelial neoplasms, mutations in PIK3CA have been occasionally reported in borderline and malignant phyllodes tumors. In contrast, mutations in MED12 are common and recurrent across the entire spectrum of benign and malignant breast fibroepithelial tumors, and are enriched in borderline/malignant phyllodes cases that still have benign fibroadenoma-like areas. In the current study, we sought to define the histologic and molecular features of PIK3CA-mutant phyllodes tumors. Methods: From 2014 to 2019, we analyzed clinical tumor samples using comprehensive genomic profiling by a hybrid capture-based DNA sequencing platform. We searched our case archive to find breast phyllodes tumors with known or likely pathogenic alterations in PIK3CA and other known tumor-related genes. All cases were clinically advanced. We reviewed pathology reports, histopathology, and patient clinical data. Results: We identified 12 (16%) of 76 breast phyllodes tumors in our case archive as PIK3CA-mutant. Median patient age for PIK3CA-mutant phyllodes tumors was 56 years. Cases consisted of 6 primary tumors, 2 local recurrences, and 4 lung metastases. Primary tumor size measured from 38 to 220 mm (median 100 mm; mean 114 mm). Digital slides were available for histology review in 9 cases. Cases showed uniformly malignant histology, with no benign or fibroadenoma-like regions. 3 cases showed malignant heterologous elements. Compared to the rest of our breast phyllodes tumor cohort, PIK3CA-mutant cases showed significantly fewer pathogenic genomic alterations in MED12 (8% vs. 55%, p=0.0037) and TP53 (17% vs. 56%, p=0.0245), as well as a trend to fewer mutations in RB1 (0% vs. 22%, p=0.11). The other most frequently mutated genes in the PIK3CA-mutant group were TERTp (70%), CDKN2A (67%), and NF1 (50%). Conclusions: PIK3CA-mutant phyllodes tumors define a unique subset of tumors characterized by aggressive histologic features and largely lacking the MED12 mutations seen in many fibroepithelial neoplasms. These findings provide compelling rationale for comprehensive genomic profiling of advanced cases of this disease in an effort to inform therapeutic options including clinical trials of PI3K-targeted agents in this setting. Citation Format: Erik A Williams, Ethan S Sokol, Dean C Pavlick, Nikunj Shah, Julia A Elvin, Jo-Anne Vergilio, Jonathan K Killian, Nhu Ngo, Douglas Lin, Vincent A Miller, Jeffrey S Ross. PIK3CA-mutant breast phyllodes tumors show a uniformly aggressive histology and significant mutual exclusivity with MED12 mutation [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-06-07.

Abstract P4-05-18: Molecular portraits of metastatic breast cancer using tissue next-generation sequencing

Abstract Introduction: Tumor sequencing efforts in breast cancer have historically focused on primary tissue samples. With the development of biomarker-based clinical trials and FDA-approved indications for anti-neoplastic agents in metastatic breast cancer (MBC), there is a deeper interest in elucidating genomic profiles of metastatic disease. We aimed to describe mutational profiles, tumor mutational burden (TMB), and microsatellite instability (MSI) status by subtype in a cohort of patients with MBC. Methods: We conducted a retrospective analysis of genomic data from 198 patients with MBC who had next generation sequencing (NGS) of tumor tissue by the TEMPUS assay (TEMPUS, Chicago, IL). The majority of patients (n=115) were treated at Northwestern Medicine. Tumors were sequenced to a minimum of 500x depth with classification of genomic alterations as actionable mutations, pathogenic mutations without known treatment, germline alterations, variants of unknown significance, copy number alterations, and chromosomal rearrangements. TMB was defined as nonsynonymous mutations per megabase (mut/MB) sequenced. Statistical analysis of TMB by subtype was conducted using independent samples T-tests. Associations were tested through two-sided Fisher’s exact test. Results: The cohort of 198 MBC patients had a median age of 51 years (range 23-88) and 52% were white, 9% black, 4% Asian, and 36% unknown. By histology, 52% had invasive ductal carcinoma, 5% had invasive lobular carcinoma, and the remainder of cases were unknown or other. The biopsy site of sequenced tissue was primary breast site in 35% and a metastatic site in the remaining. Sixty nine percent had hormone receptor positive (HR+), 21.3% had triple negative (TN), and 17.2% had HER2-positive (HER2+) MBC. Of the patients with MSI testing (n=183), 180 were MSI-stable, 3 were MSI-equivocal, and none were MSI-high. The median TMB in the whole cohort was 1.8 mut/MB (interquartile range 0.2-2.8), with a median of 2.9 (1.6-4.2) in TN compared to 1.7 (1.0-2.3) in HR+, and 1.4 (0.2-2.5) in HER2+ MBC. The mean TMB for the whole cohort and by TN, HR+, and HER2+ subtypes were 3.0, 4.8, 2.4 and 2.4, respectively. The difference in TMB was statistically significant for TN compared to HR+ (p=0.0006) and equivocal for TN compared to HER2+ (p=0.05). The most frequently seen aberrations in HR+ MBC were PIK3CA (34%), followed by TP53 (28%), ERBB2 (23%), GATA3 (23%), PTEN (13%). In HER2+ MBC alterations were seen in ERBB2 (70%), CDK12 (47%), TP53 (40%), PIK3CA (23%), ABCC3 (16%), PTEN (16%). In TN MBC TP53 (81%), KMT2C (19%), ZFHX3 (16%), PIK3CA (16%), and NCOR2 (14%) were most frequently altered. When comparing the three disease subtypes (HR+, HER2+, TN), statistically significant associations were found with TP53 mutations, most commonly observed in TN cases followed by HER2+ cases (p&amp;lt;0.00001), and ERBB2 amplifications, most commonly observed in HER2+ cases (p&amp;lt;0.00001). PIK3CA alterations were least frequently exhibited with TN compared to HR+ and/or HER2+ tumors (p=0.01). HR+ tumors had a greater proportion of GATA3 (p &amp;lt; 0.0001), and ESR1 (p=0.034) alterations compared to HR- tumors. Conversely, HER2+ tumors had a greater proportion of CKD12 (p&amp;lt; 0.0001) amplifications compared to HER- tumors. ​ Conclusions: MBC demonstrates a rich mutational profile with variations by subtype. Although TMB is lower than in several other solid malignancies, TMB greater than 10 mut/MB was observed in each subgroup, with TN MBC enriched for a higher TMB than HR+ MBC. Citation Format: Ami N Shah, Andrew A Davis, Kristen J Carroll, Firas Wehbe, Amir Behdad, Massimo Cristofanilli. Molecular portraits of metastatic breast cancer using tissue next-generation sequencing [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-05-18.

Abstract P6-10-08: AKT1 alterations following exposure to endocrine-based therapy in patients with hormone-receptor positive (HR+) metastatic breast cancer (MBC): Clinical and functional implications

Abstract Background: The AKT1 oncogene represents a central node in the well-characterized PI3K/AKT/mTOR signal transduction pathway involved in the regulation of cellular proliferation as well as therapeutic resistance. With the emergence of multiple targeted therapies for HR+ MBC, we sought to explore the prevalence of AKT1 alterations, the spectrum of co-occurring mutations, and any potential impact on response to endocrine-based therapy, including CDK 4/6 and PIK3CA inhibitors. Methods: Presence or absence of AKT1 mutation was interrogated via cell-free DNA (cfDNA) analysis (Guardant assay) among patients with HR+/HER2- MBC receiving treatment at the Massachusetts General Hospital who underwent routine clinical cfDNA testing. Genomic sequencing results were then compared to detailed clinical annotation based upon retrospective chart review. In the laboratory, HR+/HER2- T47D breast cancer cells were transfected with AKT1, under the control of a doxycycline-inducible promoter, to induce AKT1 overexpression. Sensitivity to escalating doses of fulvestrant and palbociclib were interrogated via cell-titer-glo viability assay in vitro. Results: Among 272 patients with HR+/HER2- MBC, we identified 13 patients with AKT1 mutation (4.7%). 12/13 (92%) patients had a canonical AKT1E17K alteration, one patient (8%) had an AKT1 E49K alteration. AKT1 mutations occurred concurrently with alterations in ESR1 (31%), PIK3CA (23%), TP53 (23%), ERBB2 (15%), and RB1 (8%). All patients had been exposed to some form of anti-estrogen prior to sequencing, either via (neo)adjuvant therapy and/or in the metastatic setting; 5/13 (38%) had exposure to a CDK4/6 inhibitor prior to cfDNA sequencing. Following the identification of an AKT1 alteration, examples of rapid clinical progression on fulvestrant, CDK4/6i, and/or everolimus were identified, however the presence of an AKT1 alteration was not sufficient to predict pan-resistance to these agents, as counter-examples of clinical benefit were also identified. In an index patient with an AKT1 mutation and prior progression on letrozole and palbociclib, subsequent treatment with fulvestrant and abemaciclib did not yield clinical benefit, highlighting the potential role of AKT1 in mediating clinical resistance to endocrine therapy and CDK4/6i. In the pre-clinical model, HR+/HER2- breast cancer cells demonstrated resistance to both fulvestrant and palbociclib following upregulation of exogenous AKT1 expression. Updated clinical data with additional patients and response to regimens incorporating an anti-estrogen, a CDK4/6 inhibitor, an mTOR inhibitor, and/or a PIK3CA inhibitor will be presented at the meeting. Conclusions: Activating AKT1 mutations can be identified via targeted sequencing of cfDNA in patients with HR+/HER2- MBC, and may play an important role in mediating resistance to anti-estrogens, CDK4/6 inhibitors, and other emerging targeted therapies. Heterogeneity in response to endocrine-based therapy following the identification of an AKT1 alteration may be related to tumor-extrinsic factors, the AKT1 allelic fraction, or cooperativity between multiple resistance drivers. Further research is needed to confirm these findings and guide optimal therapeutic sequencing strategies for patients with HR+/HER2- AKT1 mutant MBC. Citation Format: Seth Wander, Pingping Mao, Maxwell Lloyd, Kailey Kowalski, Gabriela N. Johnson, Giuliana Malvarosa, Beverly Moy, Leif W. Ellisen, John Iafrate, Nikhil Wagle, Aditya Bardia. AKT1 alterations following exposure to endocrine-based therapy in patients with hormone-receptor positive (HR+) metastatic breast cancer (MBC): Clinical and functional implications [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P6-10-08.