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HDAC6 is a cytoplasmic class IIB HDAC isoenzyme that is involved in the deacetylation of cytoplasmic proteins. HDAC6 also binds ubiquitinated proteins, facilitating the formation of the aggresome, to remove misfolded proteins. Previously, HDAC6-selective inhibitors were shown to decrease immunosuppression and enhance immune function of melanoma patient T-cells demonstrating an important role for HDAC6 inhibition for cancer immunotherapy. GB-1101 is a novel, orally bioavailable, nanomolar potent inhibitor of HDAC6. GB-1101 tested in vitro did not induce direct cytotoxicity to CD4+/CD8+ or NK cells at concentrations up to 300 nM. RNA-seq on GB-1101 exposed MHC-null human MSS colon cancer cell lines reveal potent induction of MHC Class I/II genes and expression numerous cancer neoantigens. To test the activity of GB-1101 in vivo, we established large (~450mm3) 4T1-luc syngeneic tumors and initiated treatment of GB-1101 given orally once daily alone and combined with anti-PD1, anti-CTLA4, and anti-PD1/anti-CTLA4 combination. The activity of anti-PD1 in this model was minimal (TGI: 3%); GB-1101 as a single agent showed more potent anti-tumor activity (TGI: 50%) compared to anti-PD1. We observed synergistic benefit when GB-1101 was combined with anti-PD1 (Combination TGI: 78%). GB-1101 showed comparable anti-tumor activity to single agent anti-CTLA4 (TGI: 43%) but revealed significant tumor regressions of large and well-established tumors when combined with anti-CTLA4 (Combination TGI: >100%). As a single agent, GB-1101 was superior to anti-PD1+anti-CTLA4 (TGI: 7%) and induced significant tumor regressions when combined with anti-PD1+anti-CTLA4 (Triple Combination TGI: >100%). In the TC-1, HPV-positive syngeneic model, GB-1101 reprogrammed the tumor immune microenvironment reducing the number of M-MDSC and TReg cells and increase the numbers of CD4+/CD8+ and NK cells. Taken together, these data support GB-1101 as an important new targeted epigenetic immunomodulator able to revoke immune privilege to enhance the clinical activity of immune checkpoint therapies. Citation Format: Kari Kotlarczyk, Mario Sepulveda, Tong Wang, Maggie Murray, Paul Gonzales, Stephen Thomas Gately. Selective HDAC6 inhibition by GB-1101 revokes tumor immune privilege and synergizes with immune checkpoint therapies to induce tumor regressions [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1359.

HDAC3 is essential for transcriptional repression and is required for the activity of NCOR1 & SMRT co-repressors complexes. Inhibition of HDAC3 results in the suppression of genes responsible for DNA damage repair and maintenance of genomic stability. HDAC3/NCOR1 also play critical roles in tumor metabolism and immune cell biology. GB-3103 is a novel and potent inhibitor of HDAC3 with an IC50 of 0.56nM that was tested for activity against human neuroendocrine prostate (NEPC) and small cell lung cancers (SCLC). RNA-seq analysis of GB-3103 treated human tumor cells revealed induction of BRCAness, inhibition of genes from DNA repair pathways and inhibition of PI3K-AKT1, MYC and p53 signaling. GB-3103 showed potent in vitro anticancer activity against the human H660 NEPC line, with an IC50 of 28nM, and 21nM against the murine PNEC30 line and potent IC50 of 16nM against NCI-H209 human lung neuroendocrine cancer. Because of the potent induction of BRCAness we used zero interaction potency (ZIP) model to test the activity of the combination of GB-3103 with a PARP inhibitor against human H660 and murine PNEC30 cell lines. These experiments revealed synergy for the combination of PARPi with GB-3103 in NEPC. GB-3103 was subsequently tested alone and in combination with olaparib in a xenograft model of H660. GB3103 (TGI=3%) showed minimal activity at the selected dose against H660 compared to single agent olaparib (TGI=67%). However, when combined with olaparib, GB-3103 revealed potent synergy (TGI=96%, p100%, p Citation Format: Stephanie Nagl, Kendra Devore, Gregg Hirschfeld, Kari Kotlarczyk, Damaris Diaz, Tong Wang, Maggie Murray, Paul Gonzales, Stephen Thomas Gately. Inhibition of HDAC3 induces BRCAness and potent synergy with PARP inhibition in neuroendocrine prostate and small cell lung cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2607.

Hemophagocytic lymphohistiocytosis (HLH) is a rare, hyperinflammatory syndrome characterized by clinical signs and symptoms of extreme inflammation. In adults, HLH is typically a complication of infections, autoimmune diseases, and malignancies. While the disease is often fatal, classic management of HLH revolves around early diagnosis and initiation of protocolized therapy. We present a case of a previously healthy 56-year-old female who developed distributive shock requiring intubation, vasopressors, and continuous venovenous hemofiltration. In the setting of multiple infectious syndromes, severe cytopenias, and rising direct hyperbilirubinemia, her diagnosis of HLH was confirmed. Therapy was initiated with dexamethasone and two doses of reduced-intensity etoposide based on the patient’s clinical course. Over the next few weeks, she continued to improve on dexamethasone monotherapy and has maintained remission up to the present with complete resolution of her cytopenias and return of baseline renal function. Our case highlights the variability in the management of probable infection-associated HLH (IHLH) with a good patient outcome. We demonstrate the potential to treat IHLH with partial protocols and minimal chemotherapeutics.

Successful immunotherapy is dependent on the regulation of an immune response concomitant with modulation of the tumor microenvironment. To study this hypothesis, we utilized a replication-incompetent adenovirus engineered to conditionally express mouse IL-12 (Ad-RTS-mIL-12), via our RheoSwitch Therapeutic System® (RTS®) gene switch which is tightly controlled by the oral activator ligand veledimex (V) in combination with the chemotherapeutic CCNU (lomustine, alkylating nitrosourea) in a GL-261 orthotopic glioma mouse model. The effects of Ad-RTS-mIL-12+veledimex (Ad+V) alone Ad 5x109vp+V 3-30mg/m2QDx14po or concomitantly with CCNU 9-18mg/m2QDx5ip were studied. Mice without treatment succumb to disease progression by day 39 (median 22 days). Ad+V 3mg/m2 monotherapy prolonged median overall survival (mOS) to 37 days. Eighty days after immunotherapy (end of study), ~65% mice receiving Ad +V 10-30mg/m2/day survived. CCNU 9mg/m2 failed to improve mOS while 18mg/m2 increased mOS to 36 days. Ad+V 3mg/m2 combined with CCNU 9mg/m2 failed to improve mOS while Ad+V 3mg/m2+CCNU 18mg/m2 increased survival with 60% surviving at study end. Ad+V 10mg/m2+CCNU 9mg/m2 failed to increase mOS over monotherapy while Ad+V 10mg/m2+CCNU 18mg/m2 resulted in 100% survival at end of study. Ad+V 30mg/m2+CCNU (9-18mg/m2) did not further improve survival with increased clinical signs over monotherapy that recovered on discontinuance of dosing. Ad+V increased tumor IL-12 (80 pg/mg) which was ~15-times greater than that of plasma while CCNU alone or in combination with Ad+V did not affect tumor and serum cytokines. Within the tumor, Ad+V elicited a dose-related increase in cytotoxic T cells and macrophages and decrease in regulatory T cells consistent with immune-mediated anti-tumor effects. Ad+V+CCNU did not further alter the tumor immune profile over Ad+V monotherapy. In summary, the controlled local immunostimulation with IL-12 combined with CCNU, warrants further investigation in GBM.

Breast cancer remains the leading cause of cancer and the third leading cause of cancer related deaths among our population with an estimated number of 246,660 new cases and 40,450 deaths in 2016. With treatment advancements, including targeted agents such as Everolimus, a mammalian target of rapamycin (mTOR) inhibitor, survivability and quality of life continue to improve. However, with the use of these agents come adverse effects, some of which are still being characterized. Our case demonstrates recurrent episodes of gastrointestinal bleeding in a 60-year-old woman being treated with Everolimus for progressive metastatic breast cancer. On endoscopy, bleeding was secondary to erosive gastritis. Previous case reports have described bleeding due to gastric antral vascular ectasia (GAVE), which was described in two prior reported cases. In our case, bleeding also occurred on a reduced dose of Everolimus compared to what is previously reported (5 mg versus 10 mg). As a result of her gastrointestinal bleeding, she required multiple endoscopic interventions including argon plasma coagulation and multipolar heater probe to achieve hemostasis. This is the first case reported of gastrointestinal bleeding not consistent with GAVE and occurring while being on a reduced dose of Everolimus. It is important to document our case so that the Gastroenterology and Hematology communities can be educated and made aware for their patient populations on Everolimus.

Dual loss of SMARCA4/SMARCA2 ATPases of the SWItch/Sucrose Non-Fermentable (SWI/SNF) complex has been reported in small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) and other tumors. Loss of SMARCA4 is the result of inactivating mutations, and the loss of SMARCA2 results from the absence of mRNA expression. Restoration of either SMARCA4 or SMARCA2 can inhibit the growth of these cancers. We have evaluated the activity of a novel, structurally rigid, and potent, Class I/IIb HDAC inhibitor, GB-3103, against human SCCOHT and other cells lines deficient in SWI/SNF complex. GB-3103 shows potent anti-proliferative activities with low nM IC50 values against human SCCOHT lines BIN67 (51nM), COV434 (35nM) and SCCOHT-1 (293nM), and SWI/SNF-deficient rhabdoid and lung tumor lines A204 (95nM), A427 (174nM), G401 (138 nM), G402 (71nM), H522 (102nM). Treatment of human BIN67 SCCOHT cell line for 72h with GB-3103 revealed potent concentration- and time-dependent induction of SMARCA2 expression at both mRNA and protein levels. Treatment of mice bearing G401 human malignant rhabdoid tumor xenografts with GB-3103 at 5 mg/kg, QD resulted in 70% tumor growth inhibition (TGI) compared to vehicle control (P Citation Format: Tong Wang, Paul Gonzales, Kari Kotlarczyk, Myung-Ja Lee, Erin Bossert, Courtney Devore, Haiyong Han, Jessica Lang, William Hendricks, Jeffrey Trent, Stephen Gately. GB-3103, an epigenetic immunomodulator, shows potent antitumor activity against tumors harboring dual loss of SMARCA4/SMARCA2 ATPases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr LB-058.

Autologous CAR T therapeutics have recently been approved for use in B-cell malignancies. While responses have been impressive using CD19 directed CAR T cells there has been a lack of comparable success for CAR T cells directed at solid tumor antigens. In an effort to address the need for effective and durable off-the shelf therapies for both hematologic and solid tumors we have developed allogeneic CAR T cells targeting the CD70 antigen. CD70 is expressed in both hematologic malignancies as well as in solid cancers such as renal cell carcinoma (RCC), while its expression in normal tissues is restricted to a subset of lymphoid cell types. CAR-T cells expressing a CD70 targeting CAR were generated by CRISPR Cas9 genome editing. T cells from healthy donors were edited to express a CD70 CAR by knocking this construct into the concurrently knocked out TCR alpha constant region (TRAC). Loss of the TCR reduces the risk of graft versus host disease enabling an allogeneic therapeutic. CD70 CAR T cells displayed potent cell killing function in vitro against CD70 expressing lymphoid and renal cancer derived cell lines across a broad range of antigen expression levels. CD70 CAR T cells also secreted IFNg, released granzyme B and proliferated in a CD70 specific manner. Furthermore, the CD70 targeting CAR T cells were able to eliminate established ccRCC tumor xenografts in mice Citation Format: Zinkal Padalia, Ashley Porras, Mary Lee Dequeant, Jason Sagert, Thao Nguyen, Matthias John, Melanie Allen, Henia Dar, Daniel Henderson, Seshidar Police, Dakai Mu, Kelly Maeng, Elaine Huang, Sarah Spencer, Nickolaus Lorson, Paul Gonzales, Chelsea Holmquist, Gregg Hirschfeld, Jonathan A. Terrett, Demetrios Kalaitzidis. Allogeneic CRISPR engineered anti CD70 CAR T cells demonstrate potent preclinical activity against both solid and hematological cancer cells [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 2551.

Kras mutations, particularly at codons 12 and 13, are reported to occur in up to one-third of human cancer cells and are considered to be undruggable targets (Stoize et al, 2015). These mutations are especially prevalent in NSCLC and CRC. Plinabulin is a new chemical entity in Phase 3, with a multi-faceted mechanism of action, including anti-angiogenesis, inhibition of tubulin polymerization and activation of JNK, which is downstream of the Kras pathway. The combination of these vascular mechanisms together with activity downstream from Kras, suggests that plinabulin could express anti-tumor activity in the presence of Kras mutations. The present report summarizes the findings with plinabulin on tumors with Kras mutations (p.12V, p.12D, p.D153V, p.G12A, p.G12C, p.G12D, p.G12V or p.G13D) in vitro and in vivo. In vitro, plinabulin was very potent (IC50 7-33 nM) as a cytotoxic agent against CRC cell lines containing a Kras mutation at p.G13D (LoVo, HCT-15, HCT116) or a BRAF + P53 mutation (HT-29) and against multiple myeloma cell lines (Singh et al, Blood, 2010) with either Nras or p.G12A mutations (IC50 In vivo, plinabulin was tested as a single agent and in combination with SOC in the CRC Kras mutation murine models LoVo and HCT-15 and the P53/Braf mutant model HT-29, the NSCLC Kras mutation model A549, the multiple myeloma Kras mutation model MM.1S and the breast Kras mutation model MDA-MB-231. As a single agent plinabulin was at most moderately active in these models, with the greatest activity seen in the LoVo, HCT-15, and MDA-MB-231 models (TGI 21-43%); the exception was a marked single agent activity observed in the MM.1S model. However, in all cases plinabulin significantly increased the anti-tumor activity of the standard of care agents (TGI 59-84%, with several CRs and increased tolerability as compared to SOC alone). In summary, plinabulin exhibits a potent anti-tumor activity either in vitro, or in vivo as a single agent or in combination with SOC agents. This activity is observed across several tumor types and mutations, but the spectrum of activity does not extend across all tumor types (e.g. inactive in vivo in Kras mutations for pancreatic cancer or melanoma). These findings are encouraging in terms of clinical studies in Kras mutations in MM, NSCLC, CRC and breast cancers. Citation Format: George Kenneth Lloyd, Lihua Du, Gloria Lee, Jessica Dalsing-Hernandez, Kari Kotlarczyk, Paul Gonzales, Steffan Nawrocki, Jennifer Carew, Lan Huang. Activity of plinabulin in tumor models with Kras mutations. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A184.

e16556 Background: Anti-tumor activity has been predicted for glucocorticoid receptor antagonists (GRAs) for some solid organ malignanies. Org34517, the first selective GRA studied in humans, has s...

Several members of the kinesin family of microtubule motor proteins play essential roles in mitotic spindle function and are potential targets for the discovery of novel antimitotic cancer therapies. KSP, also known as HsEg5, is a kinesin that plays an essential role in formation of a bipolar mitotic spindle and is required for cell cycle progression through mitosis. We identified a potent inhibitor of KSP, CK0106023, which causes mitotic arrest and growth inhibition in several human tumor cell lines. Here we show that CK0106023 is an allosteric inhibitor of KSP motor domain ATPase with a Ki of 12 nm. Among five kinesins tested, CK0106023 was specific for KSP. In tumor-bearing mice, CK0106023 exhibited antitumor activity comparable to or exceeding that of paclitaxel and caused the formation of monopolar mitotic figures identical to those produced in cultured cells. KSP was most abundant in proliferating human tissues and was absent from cultured postmitotic neurons. These findings are the first to demonstrate the feasibility of targeting mitotic kinesins for the treatment of cancer.

ODSH, a new chemical entity, is 2-0, 3-0 desulfated heparin, a desulfated heparin analog that has been shown to bind to the receptor for advanced glycation endproducts (RAGE). This interaction prevents RAGE from binding to a principal ligand, HMGB1, as well as to other ligands including S-100 calgranulin, which attenuates the activation of RAGE. RAGE activation is known to induce resistance to chemotherapy and promote pancreatic tumor cell survival. We have shown, in vivo, an increase in response with gemcitabine when combined with ODSH. ODSH was tested alone and in combination with standard of care agent, gemcitabine, against BxPC-3 human pancreatic tumor xenograft model. Single agent ODSH and gemcitabine treatment showed an 18.8% and 16.8% decrease in tumor weight, respectively. The combination regimen of ODSH and gemcitabine increased tumor growth inhibition to 37.9%. This decrease was statistically significant (P Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3698. doi:1538-7445.AM2012-3698

Adrenocortical carcinoma (ACC) is an aggressive cancer with a 5-year survival rate of 20-30%. Mitotane is the only approved drug for the treatment of patients with ACC. It often carries significant toxicities which result in the discontinuation of treatment. There are no approved second line therapies. The increased incidence of ACC in the cancer predisposition syndrome, Li-Fraumeni, suggests the involvement of the p53 pathway in ACC pathogenesis. Our analysis of the gene expression profiles of 19 ACC samples identified dysregulation of the G2/M transition and the activity of the p53 modulator, MDM2 as important in ACC pathogenesis. Polo-like kinase-1 (PLK1) is involved in the G2/M transition and acts to promote MDM2 activity through its phosphorylation. We observed that PLK1 inhibition by siRNA results in up to a 70% reduction in viability in the ACC cell lines SW-13 and H295R. Therefore, we studied PLK1 inhibition as a potential therapeutic strategy. We used the small molecule inhibitor, BI-2536, to inhibit PLK1 function in SW-13 and H295R. Drug-dose response curves demonstrated that both cell lines are sensitive to pharmacological inhibition of PLK1 (IC50 doses of 0.0094815 μM and 0.062805 μM for SW-13 and H295R, respectively.) Murine xenograft studies demonstrated that BI-2536 resulted in a statistically significant reduction in tumor growth in SW-13 but not H295R. Examination of p53 protein levels in the presence of the drug showed a dose-dependent reduction in p53 levels in SW-13, which carries a homozygous p53 mutation. The same was not true in H295R, which is p53 wild-type. To test the hypothesis that BI-2536 was decreasing mutant p53 levels by promoting its proteasomal degradation, both cell lines were treated with previously determined inhibitory concentrations of BI-2536, either alone or in combination with the proteasome inhibitor, MG132. Western blot analysis showed recovery of p53 protein when cells were concomitantly treated with MG132, supporting a role for BI-2536 as a regulator of proteasomal degradation. Both ACC cell lines are relatively insensitive to MDM2 inhibition by nutlin-3 (NCI60 GI50 values range from ∼4 uM to ∼2 uM versus ACC cell lines @13-15uM), possibly because PLK1 stimulates MDM2 and renders nutlin-3 ineffective. We therefore assessed the ability of BI-2536 to sensitize ACC cell lines to the effects of the MDM2 inhibitor. BI-2636 did show synergy with nutlin-3, shifting the IC50 in SW-13 from 19.78 μM to 6.45 μM and from 12.75 μM to 2.84 μM in H295R. These results demonstrate that inhibition of PLK1 alone or in combination with an MDM2 inhibitor warrants further investigation as a treatment for patients with ACC. Cells with mutant p53 are more sensitive to the growth inhibitory effects of BI-2536. In the context of wild-type p53, the combined inhibition of both PLK1 and MDM2 results in loss of viability. Further clinical development of PLK1 as a target should take p53 mutation status into account. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 978. doi:1538-7445.AM2012-978

22070 Background: Adrenocortical carcinoma (ACC) is a rare and lethal cancer with an average incidence of 0.7 in 1 million. Response to current standard therapy including mitotane is approximately ...

10572 Background: Adrenocortical carcinoma (ACC) is a rare and lethal cancer with an average incidence of 1 in 1.4 million. Response to current standard therapy including mitotane is approximately 22%. Herein we use gene expression profiling to identify novel targets for therapeutic intervention. Methods: The transcriptomes of 29 ACC tumors and 4 normal adrenal glands were profiled on Affymetrix Human Genome and Agilent 22K element expression arrays in order to identify genes that could represent potential therapeutic targets. Agents known to inhibit the selected targets were tested in comparison with mitotane in two ACC cell lines (H295R and SW-13) in vitro and are currently being tested in a Scid mouse xenograft system. Results: Expression of numerous kinesin family members is dysregulated within the ACC tumor population. Of interest in the ACC cells EG5 (KIF11, KSP), a kinesin important for mitotic spindle formation, is up-regulated 5.81+ 4.42 (μ+SD) and 6.48 + 0.99 fold on the Agilent and Affymetrix platforms respectively. In addition, SPARC, an albumin binding matrix associated protein is up-regulated in ACC tumors by 3.69 + 2.37 (μ+SD) and 1.56 + 0.44 on the two platforms. The anti-microtubular drugs paclitaxel and albumin bound paclitaxel (abraxane) exhibited in vitro inhibition of H295R and SW-13 cells at IC50 concentrations of 0.33 μM and 0.0078 μM for paclitaxel and 0.35 μM and 0.0087 μM for Abraxane compared to mitotane concentrations of 15.9 μM and 46.4 μM. Treatment effects of monastrol, paclitaxel, and Abraxane are currently being tested in xenograft murine models. Testing of the EG5 inhibitor S-trityl-L- cysteine is ongoing in the in vitro system. Conclusions: KIF11 inhibitors have been proposed in the treatment of cancer. These compounds may offer better anti-mitotic activity than other tubulin interacting agents with less neurotoxic side effects. Based on biological insight garnered through expression profiling of ACC tumors, the current ongoing research is investigating the use of abraxane and EG5 inhibitors in the in vitro and in vivo treatment of ACC. No significant financial relationships to disclose.