Copland, John A.

Anaplastic thyroid cancer (ATC) is one of the most lethal malignancies with a median survival time of about 4 months. Currently, there is no effective treatment, and the development of new therapies is an important and urgent issue for ATC patients. YM155 is a small molecule that was identified as the top candidate in a high-throughput screen of small molecule inhibitors performed against a panel of ATC cell lines by the National Cancer Institute. However, there were no follow-up studies investigating YM155 in ATC. Here, we determined the effects of YM155 on ATC and human primary benign thyroid cell (PBTC) survival with alamarBlue assay. Our data show that YM155 inhibited proliferation of ATC cell lines while sparing normal thyroid cells, suggesting a high therapeutic window. YM155-induced DNA damage was detected by measuring phosphorylation of γ-H2AX as a marker for DNA double-strand breaks. The formamidopyrimidine-DNA glycosylase (FPG)-modified alkaline comet assay in conjunction with reactive oxygen species (ROS) assay and glutathione (GSH)/glutathione (GSSG) assay suggests that YM155-mediated oxidative stress contributes to DNA damage. In addition, we provide evidence that YM155 causes cell cycle arrest in S phase and in the G2/M transition and causes apoptosis, as seen with flow cytometry. In this study, we show for the first time the multiple effects of YM155 in ATC cells, furthering a potential therapeutic approach for ATC.

Abstract Background: Patient derived xenografts are a standard tool used to study cancer biology and evaluate drug response phenotypes. It is well known that PDX take rate is associated with more aggressive clinical features [higher grade, biologic subtype (e.g. TNBC vs luminal)]. Previous data from a small retrospective cohort of patients (pts) with no prior chemo (n=24) suggested that primary breast tumor PDX take rate was associated with reduced overall survival. (Derose Nat Med. 2011) However, whether take rate is prognostic in newly diagnosed pts receiving standard of care neoadjuvant chemotherapy (NAC) is unknown. From the prospective BEAUTY clinical trial of pts treated with NAC in which PDX was attempted from pre-NAC as well as after chemo (post-NAC), we recently reported no difference in take rate comparing pts with or without a pathologic complete response (pCR) (Yu Breast Canc Res 2018). Herein we report the association between PDX take rate and recurrence from tumor samples implanted pre-NAC as well as post-NAC (time of surgery). Methods: The BEAUTY study is a prospective NAC study which enrolled breast cancer pts (Stage I-III; n=140) treated with neoadjuvant weekly taxane +/-trastuzumab followed by anthracycline-based chemotherapy. Percutaneous tumor biopsies were obtained prior to NAC and tumor samples from residual disease at surgery were additionally used to establish PDXs. Tumor take rate was defined as percent of pts with the development of at least one stably transplantable (passed at least for four generations) xenograft that was pathologically confirmed as breast cancer. Time to breast cancer recurrence was defined as the time from surgery to documentation of a local, regional, or distant recurrence. Gray’s test was used to assess whether the cumulative incidence (CI) of a breast cancer recurrence differs with respect to either pre-NAC PDX take or post-NAC PDX take. Results: Of 140 pts enrolled in the BEAUTY study, tumor tissue for PDX from the pre-NAC tumor was available for implantation in 113. As previously published, PDX take rate from pre-NAC tumor was 27.4% (31/113), and varied according to tumor clinical subtype [51.3% (20/39) in triple negative breast cancer (TNBC), 26.5% (9/34) in HER2+, 5.0% (2/40) in Luminal]. With median follow up of 5.7 years (range: 3 months to 6.75 years), 17 pts developed local, regional or distant disease relapse (4 of whom had a PDX established pre-NAC). The cumulative incidence of breast cancer relapse after surgery was not found to differ according to pre-NAC PDX take (5 yr CI rate: 13.6% no take; 13.4% take; p=0.8911). In pts with TNBC, the group with the highest take rate, there was also no significant difference in incidence of BC recurrence between those without PDX take and those with PDX take (5 yr CI rate: 21.4% vs 16.2% p=0.7314). PDXs were established from residual tissue from surgery (post-NAC) in 6 of 34 pts (17.6%), [specifically, TNBC: 5/9; Her2+: 1/8; and Luminal: 0/17]. Nine of these 34 pts developed a local, regional or distant recurrence. There was a tendency for pts who had a PDX established from their residual disease to have a higher incidence of a breast event (p=0.1092). The 5 year cumulative incidence of a breast event was 19.6% for pts whose post-NAC PDX did not take and 50.0% for pts whose post-NAC PDX took. Discussion: In pts receiving NAC for breast cancer, we observed no significant difference between establishment of a pre-NAC PDX and breast cancer relapse. In contrast, post-NAC PDX take rate (from residual tumor obtained from the breast at surgery) was associated with a tendency to have a higher incidence of a breast cancer event. PDXs remain a valuable tool for the evaluation of tumor biology and development of new therapeutics, especially those models established from pts with chemotherapy resistance. Citation Format: Judy C. Boughey, Vera J. Suman, Jia Yu, Katelyn Santo, Jason P. Sinnwell, Jodi M. Carter, Krishna R. Kalari, Xiaojia Tang, Sarah A. McLaughlin, Alvaro Moreno Aspitia, John A. Copland, III, Donald W. Northfelt, Richard J. Gray, Katie N. Hunt, Amy Lynn Conners, Richard Weinshilboum, Liewei Wang, Matthew P. Goetz. Association between patient derived xenograft (PDX) take rate and breast cancer recurrence in the prospective breast cancer genome guided therapy study (BEAUTY) [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD7-04.

Background: Bone metastatic prostate cancer (BMPCa), despite the initial responsiveness to androgen deprivation therapy (ADT), inevitably becomes resistant. Recent clinical trials with upfront treatment of ADT combined with chemotherapy or novel hormonal therapies (NHTs) have extended overall patient survival. These results indicate that there is significant potential for the optimization of standard-of-care therapies to delay the emergence of progressive metastatic disease. Methods: Here, we used data extracted from human bone metastatic biopsies pre- and post-abiraterone acetate/prednisone to generate a mathematical model of bone metastatic prostate cancer that can unravel the treatment impact on disease progression. Intra-tumor heterogeneity in regard to ADT and chemotherapy resistance was derived from biopsy data at a cellular level, permitting the model to track the dynamics of resistant phenotypes in response to treatment from biological first-principles without relying on data fitting. These cellular data were mathematically correlated with a clinical proxy for tumor burden, utilizing prostate-specific antigen (PSA) production as an example. Results: Using this correlation, our model recapitulated the individual patient response to applied treatments in a separate and independent cohort of patients (n = 24), and was able to estimate the initial resistance to the ADT of each patient. Combined with an intervention-decision algorithm informed by patient-specific prediction of initial resistance, we propose to optimize the sequence of treatments for each patient with the goal of delaying the evolution of resistant disease and limit cancer cell growth, offering evidence for an improvement against retrospective data. Conclusions: Our results show how minimal but widely available patient information can be used to model and track the progression of BMPCa in real time, offering a clinically relevant insight into the patient-specific evolutionary dynamics of the disease and suggesting new therapeutic options for intervention. Trial registration: NCT # 01953640. Funding: Funded by an NCI U01 (NCI) U01CA202958-01 and a Moffitt Team Science Award. CCL and DB were partly funded by an NCI PSON U01 (U01CA244101). AA was partly funded by a Department of Defense Prostate Cancer Research Program (W81XWH-15-1-0184) fellowship. LC was partly funded by a postdoctoral fellowship (PF-13-175-01-CSM) from the American Cancer Society.

Chloroethylagelastatin A (CEAA) is an analogue of agelastatin A (AA), a natural alkaloid derived from a marine sponge. It is under development for therapeutic use against brain tumors as it has excellent central nervous system (CNS) penetration and pre-clinical therapeutic activity against brain tumors. Recently, AA was shown to inhibit protein synthesis by binding to the ribosomal A-site. In this study, we developed a novel virtual screening platform to perform a comprehensive screening of various AA analogues showing that AA analogues with proven therapeutic activity including CEAA have significant ribosomal binding capacity whereas therapeutically inactive analogues show poor ribosomal binding and revealing structural fingerprint features essential for drug-ribosome interactions. In particular, CEAA was found to have greater ribosomal binding capacity than AA. Biological tests showed that CEAA binds the ribosome and contributes to protein synthesis inhibition. Our findings suggest that CEAA may possess ribosomal inhibitor activity and that our virtual screening platform may be a useful tool in discovery and development of novel ribosomal inhibitors.

AimsRecently, a novel isoform of anaplastic lymphoma kinase, with alternative transcription initiation (ALKATI), has been described in melanoma and is susceptible to targeted ALK‐inhibitor therapy. Clinical outcomes of patients with ALKATI mutated melanoma as well as correlation with immunohistochemical (IHC) methods have not yet been described.Methods and resultsClinicopathological characteristics were abstracted for 324 patients with metastatic melanoma (MM). IHC, fluorescence in‐situ hybridisation and RNA‐based digital molecular analysis assays were performed on archival tissue from 173 stage III and 192 stage IV tumours. ALKATI was identified in 12.7 and 4.8% stage III and IV tumours, respectively. Discrete presentations of the ALKATI are seen: isolated ALKATI (n = 20) and mixed ALKATI (combined ALKATI and ALKWT; n = 7). Isolated ALKWT expression (n = 4) was seen with no ALK fusions. Stage III patients showed improved survival with ALKATI expression compared to those with ALKWT or no expression [5‐year survival 80, 95% confidence interval (CI) = 57–100% versus 43%, 95% CI = 34–55%, P = 0.013]. Clinicopathological characteristics were not statistically significant. Strong diffuse cytoplasmic staining of ALK IHC (n = 12) has a sensitivity of 52.2%, specificity 100%, PPV of 100% and NPV of 92.5% of detecting isolated ALKATI.ConclusionPresence of ALKATI is a good prognostic indicator in MM. ALK IHC and digital molecular analysis can be incorporated into MM evaluation to identify patients with ALKATI for targeted therapy.

AbstractClear‐cell renal cell carcinoma (ccRCC) presents challenges to clinical management because of late‐stage detection, treatment resistance, and frequent disease recurrence. Metabolically, ccRCC has a well‐described Warburg effect utilization of glucose, but how this affects complex carbohydrate synthesis and alterations to protein and cell surface glycosylation is poorly defined. Using an imaging mass spectrometry approach, N‐glycosylation patterns and compositional differences were assessed between tumor and nontumor regions of formalin‐fixed clinical ccRCC specimens and tissue microarrays. Regions of normal kidney tissue samples were also evaluated for N‐linked glycan‐based distinctions between cortex, medullar, glomeruli, and proximal tubule features. Most notable was the proximal tubule localized detection of abundant multiantennary N‐glycans with bisecting N‐acetylglucosamine and multziple fucose residues. These glycans are absent in ccRCC tissues, while multiple tumor‐specific N‐glycans were detected with tri‐ and tetra‐antennary structures and varying levels of fucosylation and sialylation. A polycystic kidney disease tissue was also characterized for N‐glycan composition, with specific nonfucosylated glycans detected in the cyst fluid regions. Complementary to the imaging mass spectrometry analyses was an assessment of transcriptomic gene array data focused on the fucosyltransferase gene family and other glycosyltransferase genes. The transcript levels of the FUT3 and FUT6 genes responsible for the enzymes that add fucose to N‐glycan antennae were significantly decreased in all ccRCC tissues relative to matching nontumor tissues. These striking differences in glycosylation associated with ccRCC could lead to new mechanistic insight into the glycobiology underpinning kidney malignancies and suggest the potential for new therapeutic interventions and diagnostic markers.

Abstract Quiescin sulfhydryl oxidase 1 (QSOX1) is an enzyme overexpressed by many different tumor types. QSOX1 catalyzes the formation of disulfide bonds in proteins. Because short hairpin knockdowns (KD) of QSOX1 have been shown to suppress tumor growth and invasion in vitro and in vivo, we hypothesized that chemical compounds inhibiting QSOX1 enzymatic activity would also suppress tumor growth, invasion, and metastasis. High throughput screening using a QSOX1-based enzymatic assay revealed multiple potential QSOX1 inhibitors. One of the inhibitors, known as “SBI-183,” suppresses tumor cell growth in a Matrigel-based spheroid assay and inhibits invasion in a modified Boyden chamber, but does not affect viability of nonmalignant cells. Oral administration of SBI-183 inhibits tumor growth in 2 independent human xenograft mouse models of renal cell carcinoma. We conclude that SBI-183 warrants further exploration as a useful tool for understanding QSOX1 biology and as a potential novel anticancer agent in tumors that overexpress QSOX1.

122 Background: Mixed response (MR) to systemic therapy is common and associated with a poor prognosis. MR often represents a therapeutic dilemma resulting in the discontinuation of a therapy to which a portion of the tumor is still sensitive, due to the emergence of resistance in other metastatic deposits. We hypothesize that discordant responses in patients receiving immunotherapy may be associated with genomic variation within discordant tumors. Methods: We defined MR with the following criteria: 1) At least one tumor has increased in size while another tumor has decreased; 2) One or more tumors have remained stable while another has increased; 3) One or more tumors have remained stable while another has decreased. 4) A new tumor has developed while other tumors have decreased or remained stable. Between 2012 and 2017, we identified 112 patients with a MR. Two patients with metastatic lung cancer, treated with nivolumab underwent paired, simultaneous sampling of discordant tumors. Results: Both patients treated with nivolumab demonstrated genomic heterogeneity between discordant tumors on next-generation sequencing, including one patient with SCLC. Conclusions: MR remains a common and challenging therapeutic quandary. Genomic heterogeneity and differences in TMB of metastatic tumors demonstrating a MR, may be factors that contribute to this phenomenon. Further characterization of genomic discrepancies in MR may provide insights into mechanisms of emerging resistance, as well as, more rational and effective combinatorial therapies.[Table: see text]

5510 Background: Single agent aromatase inhibitor (AI) therapy is associated with limited clinical activity in ovarian cancer (OC) and endometrial cancers (EC). AI therapy was associated with a progression free survival (PFS) at 12 weeks of only 20% in relapsed OC (Bowman et al, 2002) and a median PFS of 1 month in relapsed EC (Rose et al, 2000). In Estrogen Receptor (ER) positive metastatic breast cancer, clinical studies had shown a significant prolongation of PFS with the addition of the cyclin kinase 4/6 inhibitor ribociclib to AI (Hortobagyi et al, 2016). Here, we report the results of a phase 2 clinical trial of the combination of ribociclib and letrozole in patients with relapsed ER positive OC or EC. Objectives: Primary endpoint was the proportion of patients with relapsed ER positive OC or EC alive and progression-free after 12 weeks of therapy (PFS12) with the combination of ribociclib given at a dose of 400 mg orally daily and letrozole 2.5 mg orally daily. A PFS of 45% was considered a favorable result based on the data referenced above from Bowman et al. Methods: Eligibility criteria included patients with relapsed ER positive OC or EC, with measurable disease, not previously treated with ribociclib or AIs. Xenografts were created from CT guided tumor biopsies at baseline to assess feasibility. Results: A total of 40 patients were enrolled (20 with OC and 20 with EC) ) with a median age of 61 years (range: 30-82) and 64.5 (range: 52-75) in the OC and EC groups respectively. Among the OC patients, 17 had high grade serous carcinomas and 3 had low grade serous carcinomas. 11 EC patients had endometrioid cancers (3 with grade 1 tumors) and 9 had high grade serous tumors. Ten out of 20 OC patients and 11/20 EC patients were alive and progression-free at 12 weeks (PFS12 of 50 and 55%, respectively). The most common grade 3 or higher adverse events (occurring in at least 5 pts) were leukopenia (18%), lymphopenia (18%), neutropenia (13%), and fatigue (13%). 34 tumor biopsies were suitable for injection into mice and 44% engrafted. ER expression persisted through multiple passages in mice. Two of three EC PDX models exhibited improved PFS with letrozole/ribociclib compared to letrozole alone. Conclusions: The combination of ribociclib and letrozole is associated with a promising 50% and 55% PFS12 in patients with ER positive relapsed OC or EC respectively. Creation of xenograft tumor models from CT guided biopsies of OC and EC tumors was feasible. Clinical trial information: NCT02657928.