SARC025 arms 1 and 2: A phase 1 study of the poly(ADP-ribose) polymerase inhibitor niraparib with temozolomide or irinotecan in patients with advanced Ewing sarcoma.

BACKGROUND: In preclinical Ewing sarcoma (ES) models, poly(adenosine diphosphate ribose) polymerase (PARP) inhibitors were identified as a potential therapeutic strategy with synergy in combination with cytotoxic agents. This study evaluated the safety and dosing of the PARP1/2 inhibitor niraparib (NIR) with temozolomide (TMZ; arm 1) or irinotecan (IRN; arm 2) in patients with pretreated ES. METHODS: Eligible patients in arm 1 received continuous NIR daily and escalating TMZ (days 2-6 [D2-6]) in cohort A. Subsequent patients received intermittent NIR dosing (cohort B), with TMZ re-escalation in cohort C. In arm 2, patients were assigned to NIR (days 1-7 [D1-7]) and escalating doses of IRN (D2-6). RESULTS: From July 2014 to May 2018, 29 eligible patients (23 males and 6 females) were enrolled in arms 1 and 2, which had 7 dose levels combined. Five patients experienced at least 1 dose-limiting toxicity (DLT) in arm 1 (grade 4 [G4] neutropenia for >7 days or G4 thrombocytopenia), and 3 patients experienced at least 1 DLT in arm 2 (grade 3 [G3] colitis, G3 anorexia, or G3 alanine aminotransferase elevation). The maximum tolerated dose was NIR at 200 mg every day on D1-7 plus TMZ at 30 mg/m2 every day on D2-6 (arm 1) or NIR at 100 mg every day on D1-7 plus IRN at 20 mg/m2 every day on D2-6 (arm 2). One confirmed partial response was observed in arm 2; the median progression-free survival was 9.0 weeks (95% CI, 7.0-10.1 weeks) and 16.3 weeks (95% CI, 5.1-69.7 weeks) in arms 1 and 2, respectively. The median decrease in tumor poly(ADP-ribose) activity was 89% (range, 83%-98%). CONCLUSIONS: The combination of NIR and TMZ or IRN was tolerable, but at lower doses in comparison with conventional cytotoxic combinations. A triple-combination study of NIR, IRN, and TMZ has commenced.

NADPH oxidase 5 (NOX5)-induced reactive oxygen signaling modulates normoxic HIF-1α and p27Kip1 expression in malignant melanoma and other human tumors.

NADPH oxidase 5 (NOX5) generated reactive oxygen species (ROS) have been implicated in signaling cascades that regulate cancer cell proliferation. To evaluate and validate NOX5 expression in human tumors, we screened a broad range of tissue microarrays (TMAs), and report substantial overexpression of NOX5 in malignant melanoma and cancers of the prostate, breast, and ovary. In human UACC-257 melanoma cells that possesses high levels of functional endogenous NOX5, overexpression of NOX5 resulted in enhanced cell growth, increased numbers of BrdU positive cells, and increased γ-H2AX levels. Additionally, NOX5-overexpressing (stable and inducible) UACC-257 cells demonstrated increased normoxic HIF-1α expression and decreased p27Kip1 expression. Similarly, increased normoxic HIF-1α expression and decreased p27Kip1 expression were observed in stable NOX5-overexpressing clones of KARPAS 299 human lymphoma cells and in the human prostate cancer cell line, PC-3. Conversely, knockdown of endogenous NOX5 in UACC-257 cells resulted in decreased cell growth, decreased HIF-1α expression, and increased p27Kip1 expression. Likewise, in an additional human melanoma cell line, WM852, and in PC-3 cells, transient knockdown of endogenous NOX5 resulted in increased p27Kip1 and decreased HIF-1α expression. Knockdown of endogenous NOX5 in UACC-257 cells resulted in decreased Akt and GSK3ß phosphorylation, signaling pathways known to modulate p27Kip1 levels. In summary, our findings suggest that NOX5 expression in human UACC-257 melanoma cells could contribute to cell proliferation due, in part, to the generation of high local concentrations of extracellular ROS that modulate multiple pathways that regulate HIF-1α and networks that signal through Akt/GSK3ß/p27Kip1 .

Randomized phase II trial of cyclophosphamide and the oral poly (ADP-ribose) polymerase inhibitor veliparib in patients with recurrent, advanced triple-negative breast cancer.

Background In tumors carrying BRCA mutations, DNA damage caused by standard cytotoxic chemotherapy can be potentiated by poly [ADP-ribose] polymerase (PARP) inhibitors, leading to increased cell death through synthetic lethality. Individuals carrying mutations in BRCA have an increased incidence of triple negative breast cancer (TNBC). In order to assess the role of PARP inhibition in the treatment of TNBC, we conducted a randomized phase II trial of the combination of veliparib, a small molecule PARP inhibitor, with the cytotoxic agent cyclophosphamide versus cyclophosphamide alone in patients with refractory TNBC. Methods Adult patients with TNBC were randomized to receive oral cyclophosphamide 50 mg once daily with or without oral veliparib at 60 mg daily in 21-day cycles. Patients on the cyclophosphamide arm could crossover to the combination arm at disease progression. Results Forty-five patients were enrolled; 18 received cyclophosphamide alone and 21 received the combination as their initial treatment regimen. Lymphopenia was the most common grade 3/4 toxicity noted in both arms. One patient in the cyclophosphamide alone arm, and 2 in the combination arm had objective responses. Response rates and median progression free survival did not significantly differ between both treatment arms. Conclusion The addition of veliparib to cyclophosphamide, at the dose and schedule evaluated, did not improve the response rate over cyclophosphamide treatment alone in patients with heavily pre-treated triple-negative breast cancer.

Characterization of Batracylin-induced Renal and Bladder Toxicity in Rats.

Batracylin (NSC-320846) is a dual inhibitor of DNA topoisomerases I and II. Batracylin advanced as an anticancer agent to Phase I clinical trials where dose limiting hemorrhagic cystitis (bladder inflammation and bleeding) was observed. To further investigate batracylin's mechanism of toxicity, studies were conducted in Fischer 344 rats. Once daily oral administration of 16 or 32 mg/kg batracylin to rats for 4 days caused overt toxicity. Abnormal clinical observations and adverse effects on clinical pathology, urinalysis, and histology indicated acute renal damage and urothelial damage and bone marrow dysfunction. Scanning electron microscopy revealed sloughing of the superficial and intermediate urothelial layers. DNA damage was evident in kidney and bone marrow as indicated by histone γ-H2AX immunofluorescence. After a single oral administration of 16 or 32 mg/kg, the majority of batracylin was converted to N-acetylbatracylin (NAB) with a half-life of 4 hr to 11 hr. Mesna (Mesnex™), a drug known to reduce the incidence of hemorrhagic cystitis induced by ifosfamide or cyclophosphamide, was administered to rats prior to batracylin, but did not alleviate batracylin-induced bladder and renal toxicity. These findings suggest that batracylin results in DNA damage-based mechanisms of toxicity and not an acrolein-based mechanism of toxicity as occurs after ifosfamide or cyclophosphamide administration.

Rationale for poly(ADP-ribose) polymerase (PARP) inhibitors in combination therapy with camptothecins or temozolomide based on PARP trapping versus catalytic inhibition.

We recently showed that poly(ADP-ribose) polymerase (PARP) inhibitors exert their cytotoxicity primarily by trapping PARP-DNA complexes in addition to their NAD(+)-competitive catalytic inhibitory mechanism. PARP trapping is drug-specific, with olaparib exhibiting a greater ability than veliparib, whereas both compounds are potent catalytic PARP inhibitors. Here, we evaluated the combination of olaparib or veliparib with therapeutically relevant DNA-targeted drugs, including the topoisomerase I inhibitor camptothecin, the alkylating agent temozolomide, the cross-linking agent cisplatin, and the topoisomerase II inhibitor etoposide at the cellular and molecular levels. We determined PARP-DNA trapping and catalytic PARP inhibition in genetically modified chicken lymphoma DT40, human prostate DU145, and glioblastoma SF295 cancer cells. For camptothecin, both PARP inhibitors showed highly synergistic effects due to catalytic PARP inhibition, indicating the value of combining either veliparib or olaparib with topoisomerase I inhibitors. On the other hand, for temozolomide, PARP trapping was critical in addition to catalytic inhibition, consistent with the fact that olaparib was more effective than veliparib in combination with temozolomide. For cisplatin and etoposide, olaparib only showed no or a weak combination effect, which is consistent with the lack of involvement of PARP in the repair of cisplatin- and etoposide-induced lesions. Hence, we conclude that catalytic PARP inhibitors are highly effective in combination with camptothecins, whereas PARP inhibitors capable of PARP trapping are more effective with temozolomide. Our study provides insights in combination treatment rationales for different PARP inhibitors.

Pharmacodynamic effects of the PARP inhibitor talazoparib (MDV3800, BMN 673) in patients with BRCA-mutated advanced solid tumors.

PURPOSE: Talazoparib is an inhibitor of the poly (ADP-ribose) polymerase (PARP) family of enzymes and is FDA-approved for patients with (suspected) deleterious germline BRCA1/2-mutated, HER2­negative, locally advanced or metastatic breast cancer. Because knowledge of the pharmacodynamic (PD) effects of talazoparib in patients has been limited to studies of PARP enzymatic activity (PARylation) in peripheral blood mononuclear cells, we developed a study to assess tumoral PD response to talazoparib treatment (NCT01989546). METHODS: We administered single-agent talazoparib (1 mg/day) orally in 28-day cycles to adult patients with advanced solid tumors harboring (suspected) deleterious BRCA1 or BRCA2 mutations. The primary objective was to examine the PD effects of talazoparib; the secondary objective was to determine overall response rate (ORR). Tumor biopsies were mandatory at baseline and post-treatment on day 8 (optional at disease progression). Biopsies were analyzed for PARylation, DNA damage response (γH2AX), and epithelial‒mesenchymal transition. RESULTS: Nine patients enrolled in this trial. Four of six patients (67%) evaluable for the primary PD endpoint exhibited a nuclear γH2AX response on day 8 of treatment, and five of six (83%) also exhibited strong suppression of PARylation. A transition towards a more mesenchymal phenotype was seen in 4 of 6 carcinoma patients, but this biological change did not affect γH2AX or PAR responses. The ORR was 55% with the five partial responses lasting a median of six cycles. CONCLUSION: Intra-tumoral DNA damage response and inhibition of PARP enzymatic activity were confirmed in patients with advanced solid tumors harboring BRCA1/2 mutations after 8 days of talazoparib treatment.

Inhibition of NAD+-Dependent Metabolic Processes Induces Cellular Necrosis and Tumor Regression in Rhabdomyosarcoma Models.

PURPOSE: Deregulated metabolism in cancer cells represents a vulnerability that may be therapeutically exploited to benefit patients. One such target is nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in the NAD+ salvage pathway. NAMPT is necessary for efficient NAD+ production and may be exploited in cells with increased metabolic demands. We have identified NAMPT as a dependency in rhabdomyosarcoma (RMS), a malignancy for which novel therapies are critically needed. Here we describe the effect of NAMPT inhibition on RMS proliferation and metabolism in vitro and in vivo. EXPERIMENTAL DESIGN: Assays of proliferation and cell death were used to determine the effects of pharmacologic NAMPT inhibition in a panel of ten molecularly diverse RMS cell lines. Mechanism of the clinical NAMPTi OT-82 was determined using measures of NAD+ and downstream NAD+-dependent functions, including energy metabolism. We used orthotopic xenograft models to examine tolerability, efficacy, and drug mechanism in vivo. RESULTS: Across all ten RMS cell lines, OT-82 depleted NAD+ and inhibited cell growth at concentrations ≤1 nmol/L. Significant impairment of glycolysis was a universal finding, with some cell lines also exhibiting diminished oxidative phosphorylation. Most cell lines experienced profound depletion of ATP with subsequent irreversible necrotic cell death. Importantly, loss of NAD and glycolytic activity were confirmed in orthotopic in vivo models, which exhibited complete tumor regressions with OT-82 treatment delivered on the clinical schedule. CONCLUSIONS: RMS is highly vulnerable to NAMPT inhibition. These findings underscore the need for further clinical study of this class of agents for this malignancy.

A phase I study of veliparib with cyclophosphamide and veliparib combined with doxorubicin and cyclophosphamide in advanced malignancies.

PURPOSE: Veliparib (V), an oral poly(ADP-ribose) polymerase (PARP) inhibitor, potentiates effects of alkylating agents and topoisomerase inhibitors in preclinical tumor models. We conducted a phase I trial of V with iv cyclophosphamide (C) and V plus iv doxorubicin (A) and C. METHODS: Objectives were to establish the maximum tolerated dose (MTD) of the combinations, characterize V pharmacokinetics (PK) in the presence and absence of C, measure PAR in peripheral blood mononuclear cells (PBMCs) and γH2AX in circulating tumor cells (CTCs). In Group 1, dose escalations of V from 10 to 50 mg every 12 h Days 1-4 plus C 450 to 750 mg/m2 Day 3 in 21-day cycles were evaluated. In Group 2, V doses ranged from 50 to 150 mg every 12 h Days 1-4 with AC (60/600 mg/m2) Day 3 in 21-day cycles. In Group 3, patients received AC Day 1 plus V Days 1-7, and in Group 4, AC Day 1 plus V Days 1-14 was given in 21-day cycles to evaluate effects on γH2AX foci. RESULTS: Eighty patients were enrolled. MTD was not reached for V and C. MTD for V and AC was V 100 mg every 12 h Days 1-4 with AC (60/600 mg/m2) Day 3 every 21 days. V PK appears to be dose-dependent and has no effect on the PK of C. Overall, neutropenia and anemia were the most common adverse events. Objective response in V and AC treated groups was 22% (11/49). Overall clinical benefit rate was 31% (25/80). PAR decreased in PBMCs. Percentage of γH2AX-positive CTCs increased after treatment with V and AC. CONCLUSION: V and AC can be safely combined. Activity was observed in patients with metastatic breast cancer.

A phase 1 and pharmacodynamic study of chronically-dosed, single-agent veliparib (ABT-888) in patients with BRCA1- or BRCA2-mutated cancer or platinum-refractory ovarian or triple-negative breast cancer.

PURPOSE: BRCA1 or BRCA2 mutated cancers (BRCAmut) have intrinsic sensitivity to PARP inhibitors due to deficiency in homologous recombination-mediated DNA repair. There are similarities between BRCAmut and BRCAwt ovarian and basal-like breast cancers. This phase I study determined the recommended phase II dose (RP2D) and preliminary efficacy of the PARP inhibitor, veliparib (ABT-888), in these patients. PATIENTS AND METHODS: Patients (n = 98) were dosed with veliparib 50-500 mg twice daily (BID). The BRCAmut cohort (n = 70) contained predominantly ovarian (53%) and breast (23%) cancers; the BRCAwt cohort (n = 28) consisted primarily of breast cancer (86%). The MTD, DLT, adverse events, PK, PD, and clinical response were assessed. RESULTS: DLTs were grade 3 nausea/vomiting at 400 mg BID in a BRCAmut carrier, grade 2 seizure at 400 mg BID in a patient with BRCAwt cancer, and grade 2 seizure at 500 mg BID in a BRCAmut carrier. Common toxicities included nausea (65%), fatigue (45%), and lymphopenia (38%). Grade 3/4 toxicities were rare (highest lymphopenia at 15%). Overall response rate (ORR) was 23% (95% CI 13-35%) in BRCAmut overall, and 37% (95% CI 21-55%) at 400 mg BID and above. In BRCAwt, ORR was 8% (95% CI 1-26%), and clinical benefit rate was 16% (95% CI 4-36%), reflecting prolonged stable disease in some patients. PK was linear with dose and was correlated with response and nausea. CONCLUSIONS: Continuous veliparib is safe and tolerable. The RP2D was 400 mg BID. There is evidence of clinical activity of veliparib in patients with BRCAmut and BRCAwt cancers.

Safety, Antitumor Activity, and Biomarker Analysis in a Phase I Trial of the Once-daily Wee1 Inhibitor Adavosertib (AZD1775) in Patients with Advanced Solid Tumors.

PURPOSE: The Wee1 kinase inhibitor adavosertib abrogates cell-cycle arrest, leading to cell death. Prior testing of twice-daily adavosertib in patients with advanced solid tumors determined the recommended phase II dose (RPh2D). Here, we report results for once-daily adavosertib. PATIENTS AND METHODS: A 3 + 3 dose-escalation design was used, with adavosertib given once daily on days 1 to 5 and 8 to 12 in 21-day cycles. Molecular biomarkers of Wee1 activity, including tyrosine 15-phosphorylated Cdk1/2 (pY15-Cdk), were assessed in paired tumor biopsies. Whole-exome sequencing and RNA sequencing of remaining tumor tissue identified potential predictive biomarkers. RESULTS: Among the 42 patients enrolled, the most common toxicities were gastrointestinal and hematologic; dose-limiting toxicities were grade 4 hematologic toxicity and grade 3 fatigue. The once-daily RPh2D was 300 mg. Six patients (14%) had confirmed partial responses: four ovarian, two endometrial. Adavosertib plasma exposures were similar to those from twice-daily dosing. On cycle 1 day 8 (pre-dose), tumor pY15-Cdk levels were higher than baseline in four of eight patients, suggesting target rebound during the day 5 to 8 dosing break. One patient who progressed rapidly had a tumor WEE1 mutation and potentially compensatory PKMYT1 overexpression. Baseline CCNE1 overexpression occurred in both of two responding patients, only one of whom had CCNE1 amplification, and in zero of three nonresponding patients. CONCLUSIONS: We determined the once-daily adavosertib RPh2D and observed activity in patients with ovarian or endometrial carcinoma, including two with baseline CCNE1 mRNA overexpression. Future studies will determine whether CCNE1 overexpression is a predictive biomarker for adavosertib.

Inhibition of nicotinamide phosphoribosyltransferase (NAMPT) with OT-82 induces DNA damage, cell death, and suppression of tumor growth in preclinical models of Ewing sarcoma.

NAMPT mediates the rate-limiting step of the NAD salvage pathway, which maintains cellular bioenergetics and provides a necessary substrate for functions essential to rapidly proliferating cancer cells. In this study, we evaluated the efficacy and mechanisms of action of OT-82, a novel, high-potency NAMPT inhibitor with a favorable toxicity profile, in preclinical models of Ewing sarcoma (EWS), an aggressive pediatric malignancy with previously reported selective sensitivity to NAMPT inhibition. We show that OT-82 decreased NAD concentration and impaired proliferation of EWS cells in a dose-dependent manner, with IC50 values in the single-digit nanomolar range. Notably, genetic depletion of NAMPT phenocopied pharmacological inhibition. On-target activity of OT-82 was confirmed with the addition of NMN, the product of NAMPT, which rescued NAD concentration and EWS cellular viability. Mechanistically, OT-82 treatment resulted in impaired DNA damage repair through loss of PARP activity, G2 cell-cycle arrest, and apoptosis in EWS cells. Additional consequences of OT-82 treatment included reduction of glycolytic and mitochondrial activity. In vivo, OT-82 impaired tumor growth and prolonged survival in mice bearing EWS xenografts. Importantly, antitumor effect correlated with pharmacodynamic markers of target engagement. Furthermore, combining low-dose OT-82 with low doses of agents augmenting DNA damage demonstrated enhanced antitumor activity in vitro and in vivo. Thus, OT-82 treatment represents a potential novel targeted approach for the clinical treatment of EWS.

Designing phase 0 cancer clinical trials.

Phase 0 trials are designed primarily to evaluate the pharmacodynamic and/or pharmacokinetic properties of selected investigational agents before initiating more traditional phase I testing. One of the major objectives of phase 0 trials is to interrogate and refine a target or biomarker assay for drug effect in human samples implementing procedures developed and validated in preclinical models. Thus, close collaboration between laboratory scientists and clinical investigators is essential to the design and conduct of phase 0 trials. Given the relatively small number of patients and tissue samples, showing a significant drug effect in phase 0 trials requires precise and reproducible assay procedures and innovative statistical methodology. Furthermore, phase 0 trials involving limited exposure of a study agent administered at low doses and/or for a short period allow them to be initiated under the Food and Drug Administration exploratory investigational new drug guidance with less preclinical toxicity data than usually required for traditional first-in-human studies. Because of the very limited drug exposure, phase 0 trials offer no chance of therapeutic benefit, which can impede patient enrollment, particularly if invasive tumor biopsies are required. The challenges to accrual are not insurmountable, however, and well-designed and executed phase 0 trials are feasible and have great potential for improving the efficiency and success of subsequent trials, particularly those evaluating molecularly targeted agents.

NCI Comparative Oncology Program Testing of Non-Camptothecin Indenoisoquinoline Topoisomerase I Inhibitors in Naturally Occurring Canine Lymphoma.

PURPOSE: Only one chemical class of topoisomerase I (TOP1) inhibitors is FDA approved, the camptothecins with irinotecan and topotecan widely used. Because of their limitations (chemical instability, drug efflux-mediated resistance, and diarrhea), novel TOP1 inhibitors are warranted. Indenoisoquinoline non-camptothecin topoisomerase I (TOP1) inhibitors overcome chemical instability and drug resistance that limit camptothecin use. Three indenoisoquinolines, LMP400 (indotecan), LMP776 (indimitecan), and LMP744, were examined in a phase I study for lymphoma-bearing dogs to evaluate differential efficacy, pharmacodynamics, toxicology, and pharmacokinetics. EXPERIMENTAL DESIGN: Eighty-four client-owned dogs with lymphomas were enrolled in dose-escalation cohorts for each indenoisoquinoline, with an expansion phase for LMP744. Efficacy, tolerability, pharmacokinetics, and target engagement were determined. RESULTS: The MTDs were 17.5 mg/m2 for LMP 776 and 100 mg/m2 for LMP744; bone marrow toxicity was dose-limiting; up to 65 mg/m2 LMP400 was well-tolerated and MTD was not reached. None of the drugs induced notable diarrhea. Sustained tumor accumulation was observed for LMP744; γH2AX induction was demonstrated in tumors 2 and 6 hours after treatment; a decrease in TOP1 protein was observed in most lymphoma samples across all compounds and dose levels, which is consistent with the fact that tumor response was also observed at low doses LMP744. Objective responses were documented for all indenoisoquinolines; efficacy (13/19 dogs) was greatest for LMP744. CONCLUSIONS: These results demonstrate proof-of-mechanism for indenoisoquinoline TOP1 inhibitors supporting their further clinical development. They also highlight the value of the NCI Comparative Oncology Program (https://ccr.cancer.gov/Comparative-Oncology-Program) for evaluating novel therapies in immunocompetent pets with cancers.

A Phase I Clinical Trial of the Poly(ADP-ribose) Polymerase Inhibitor Veliparib and Weekly Topotecan in Patients with Solid Tumors.

Purpose: To determine the dose limiting toxicities (DLT), maximum tolerated dose (MTD), and recommended phase II dose (RP2D) of veliparib in combination with weekly topotecan in patients with solid tumors. Correlative studies were included to assess the impact of topotecan and veliparib on poly(ADP-ribose) levels in peripheral blood mononuclear cells, serum pharmacokinetics of both agents, and potential association of germline repair gene mutations with outcome.Experimental Design: Eligible patients had metastatic nonhematologic malignancies with measurable disease. Using a 3 + 3 design, patients were treated with veliparib orally twice daily on days 1-3, 8-10, and 15-17 and topotecan intravenously on days 2, 9, and 16 every 28 days. Tumor responses were assessed by RECIST.Results: Of 58 patients enrolled, 51 were evaluable for the primary endpoint. The MTD and RP2D was veliparib 300 mg twice daily on days 1-3, 8-10, and 15-17 along with topotecan 3 mg/m2 on days 2, 9, and 16 of a 28-day cycle. DLTs were grade 4 neutropenia lasting >5 days. The median number of cycles was 2 (1-26). The objective response rate was 10%, with 1 complete and 4 partial responses. Twenty-two patients (42%) had stable disease ranging from 4 to 26 cycles. Patients with germline BRCA1, BRCA2, or RAD51D mutations remained on study longer than those without homologous recombination repair (HRR) gene mutations (median 4 vs. 2 cycles).Conclusions: Weekly topotecan in combination with veliparib has a manageable safety profile and appears to warrant further investigation. Clin Cancer Res; 24(4); 744-52. ©2017 AACR.

Phase I Study of ATR Inhibitor M6620 in Combination With Topotecan in Patients With Advanced Solid Tumors.

Purpose Our preclinical work identified depletion of ATR as a top candidate for topoisomerase 1 (TOP1) inhibitor synthetic lethality and showed that ATR inhibition sensitizes tumors to TOP1 inhibitors. We hypothesized that a combination of selective ATR inhibitor M6620 (previously VX-970) and topotecan, a selective TOP1 inhibitor, would be tolerable and active, particularly in tumors with high replicative stress. Patients and Methods This phase I study tested the combination of M6620 and topotecan in 3-week cycles using 3 + 3 dose escalation. The primary end point was the identification of the maximum tolerated dose of the combination. Efficacy and pharmacodynamics were secondary end points. Results Between September 2016 and February 2017, 21 patients enrolled. The combination was well tolerated, which allowed for dose escalation to the highest planned dose level (topotecan 1.25 mg/m2, days 1 to 5; M6620 210 mg/m2, days 2 and 5). One of six patients at this dose level experienced grade 4 thrombocytopenia that required transfusion, a dose-limiting toxicity. Most common treatment-related grade 3 or 4 toxicities were anemia, leukopenia, and neutropenia (19% each); lymphopenia (14%); and thrombocytopenia (10%). Two partial responses (≥ 18 months, ≥ 7 months) and seven stable disease responses ≥ 3 months (median, 9 months; range, 3 to 12 months) were seen. Three of five patients with small-cell lung cancer, all of whom had platinum-refractory disease, had a partial response or prolonged stable disease (10, ≥ 6, and ≥ 7 months). Pharmacodynamic studies showed preliminary evidence of ATR inhibition and enhanced DNA double-stranded breaks in response to the combination. Conclusion To our knowledge, this report is the first of an ATR inhibitor-chemotherapy combination. The maximum dose of topotecan plus M6620 is tolerable. The combination seems particularly active in platinum-refractory small-cell lung cancer, which tends not to respond to topotecan alone. Phase II studies with biomarker evaluation are ongoing.

Exposure-Response of Veliparib to Inform Phase II Trial Design in Refractory or Relapsed Patients with Hematological Malignancies.

Purpose: A phase I trial of veliparib in combination with topotecan plus carboplatin (T+C) demonstrated a 33% objective response rate in patients with hematological malignancies. The objective is to perform exposure-response analysis to inform the phase II trial design.Experimental Design: Pharmacokinetic, efficacy, and safety data from 95 patients, who were administered 10 to 100 mg b.i.d. doses of veliparib for either 8, 14, or 21 days with T+C, were utilized for exposure-efficacy (objective response and overall survival) and exposure-safety (≥grade 3 mucositis) analysis. Multivariate cox proportional hazards and logistic regression analyses were conducted. The covariates evaluated were disease status, duration of treatment, and number of prior therapies.Results: The odds of having objective response were 1.08-fold with 1,000 ng/hr/mL increase in AUC, 1.8-fold with >8 days treatment, 2.8-fold in patients with myeloproliferative neoplasms (MPN), and 0.5-fold with ≥2 prior therapies. Based on analysis of overall survival, hazard of death decreased by 1.5% for 1,000 ng/hr/mL increase in AUC, 39% with >8 days treatment, 44% in patients with MPN, while increased by 19% with ≥2 prior therapies. The odds of having ≥grade 3 mucositis increased by 29% with 1,000 ng.h/mL increase in AUC.Conclusions: Despite shallow exposure-efficacy relationship, doses lower than 80 mg do not exceed veliparib single agent preclinical IC50 Shallow exposure-mucositis relationship also supports the 80-mg dose. Based on benefit/risk assessment, veliparib at a dose of 80 mg b.i.d. for at least 14 days in combination with T+C is recommended to be studied in MPN patients. Clin Cancer Res; 23(21); 6421-9. ©2017 AACR.

Parent-Metabolite Pharmacokinetic Modeling and Pharmacodynamics of Veliparib (ABT-888), a PARP Inhibitor, in Patients With BRCA 1/2-Mutated Cancer or PARP-Sensitive Tumor Types.

Veliparib (ABT-888) is a novel oral poly-ADP-ribose polymerase (PARP) inhibitor that is being developed for the treatment of hematologic malignancies and solid tumors. Although the pharmacokinetics of veliparib have been studied in combination with cytotoxic agents, limited information exists regarding the pharmacokinetics (PK) of chronically dosed single-agent veliparib in patients with either BRCA 1/2-mutated cancer or PARP-sensitive tumors. The objectives of the current analysis were to characterize the population pharmacokinetics of veliparib and its primary, active metabolite, M8, and to evaluate the relationship between veliparib and M8 concentrations and poly-ADP-ribose (PAR) level observed in peripheral blood mononuclear cells (PBMCs). Seventy-one subjects contributed with veliparib plasma concentrations, M8 plasma concentrations, and PAR levels in PBMCs. Veliparib and M8 concentrations were modeled simultaneously using a population PK approach. A 2-compartment model with delayed first-order absorption and the elimination parameterized as renal (CLR /F) and nonrenal clearance (CLNR /F) adequately described veliparib pharmacokinetics. The pharmacokinetics of the M8 metabolite was described with a 2-compartment model. Creatinine clearance(CLCR ) and lean body mass (LBM) were identified as significant predictors of veliparib CLR /F and central volume of distribution, respectively. For a typical subject (LBM, 48 kg; CLCR , 95 mL/min), total clearance (CLR /F + CLNR /F), and central and peripheral volume of distribution for veliparib were estimated as 17.3 L/h, 98.7 L, and 48.3 L, respectively. At least 50% inhibition of PAR levels in PBMCs was observed at dose levels ranging from 50 to 500 mg.

Phosphorylated fraction of H2AX as a measurement for DNA damage in cancer cells and potential applications of a novel assay.

Phosphorylated H2AX (γ-H2AX) is a sensitive marker for DNA double-strand breaks (DSBs), but the variability of H2AX expression in different cell and tissue types makes it difficult to interpret the meaning of the γ-H2AX level. Furthermore, the assays commonly used for γ-H2AX detection utilize laborious and low-throughput microscopy-based methods. We describe here an ELISA assay that measures both phosphorylated H2AX and total H2AX absolute amounts to determine the percentage of γ-H2AX, providing a normalized value representative of the amount of DNA damage. We demonstrate the utility of the assay to measure DSBs introduced by either ionizing radiation or DNA-damaging agents in cultured cells and in xenograft models. Furthermore, utilizing the NCI-60 cancer cell line panel, we show a correlation between the basal fraction of γ-H2AX and cellular mutation levels. This additional application highlights the ability of the assay to measure γ-H2AX levels in many extracts at once, making it possible to correlate findings with other cellular characteristics. Overall, the γ-H2AX ELISA represents a novel approach to quantifying DNA damage, which may lead to a better understanding of mutagenic pathways in cancer and provide a useful biomarker for monitoring the effectiveness of DNA-damaging anticancer agents.

Matrix Screen Identifies Synergistic Combination of PARP Inhibitors and Nicotinamide Phosphoribosyltransferase (NAMPT) Inhibitors in Ewing Sarcoma.

Purpose: Although many cancers are showing remarkable responses to targeted therapies, pediatric sarcomas, including Ewing sarcoma, remain recalcitrant. To broaden the therapeutic landscape, we explored the in vitro response of Ewing sarcoma cell lines against a large collection of investigational and approved drugs to identify candidate combinations.Experimental Design: Drugs displaying activity as single agents were evaluated in combinatorial (matrix) format to identify highly active, synergistic drug combinations, and combinations were subsequently validated in multiple cell lines using various agents from each class. Comprehensive metabolomic and proteomic profiling was performed to better understand the mechanism underlying the synergy. Xenograft experiments were performed to determine efficacy and in vivo mechanism.Results: Several promising candidates emerged, including the combination of small-molecule PARP and nicotinamide phosphoribosyltransferase (NAMPT) inhibitors, a rational combination as NAMPTis block the rate-limiting enzyme in the production of nicotinamide adenine dinucleotide (NAD+), a necessary substrate of PARP. Mechanistic drivers of the synergistic cell killing phenotype of these combined drugs included depletion of NMN and NAD+, diminished PAR activity, increased DNA damage, and apoptosis. Combination PARPis and NAMPTis in vivo resulted in tumor regression, delayed disease progression, and increased survival.Conclusions: These studies highlight the potential of these drugs as a possible therapeutic option in treating patients with Ewing sarcoma. Clin Cancer Res; 23(23); 7301-11. ©2017 AACR.

Phase I/Ib study of olaparib and carboplatin in women with triple negative breast cancer.

PURPOSE: To investigate the safety, activity, and potential biomarkers of response to olaparib and carboplatin combination in sporadic triple negative breast cancer (TNBC). EXPERIMENTAL DESIGN: Metastatic or recurrent TNBC patients with no germline BRCA mutation or with BRCAPro scores <10% and a negative family history were eligible. A 3+3 dose escalation tested olaparib capsules (400mg bid, days1-7) with carboplatin AUC3-5 on day1 or 2 every 21 days, ≤ 8 cycles, with olaparib 400mg bid maintenance. Peripheral blood mononuclear cells were collected for polymorphisms and PAR levels, and paired tumor biopsies (pre-/post-cycle 1) for proteomics and apoptosis endpoints. RESULTS: 28 women were treated (median 5 prior regimens [0-12]). Dose-limiting toxicity was thrombocytopenia, and symptomatic hyponatremia with carboplatin AUC5. The maximum tolerated dose was olaparib 400mg bid+carboplatin AUC4. Grade 3 and 4 adverse events included neutropenia (36%), thrombocytopenia (11%), and anemia (11%). Responses included 1 complete response (CR; 69+months) and 5/27 partial responses (19%; median 4months [4-7]), for a response rate of 22%. Biomarker findings did not correlate with response. The long-term CR patient with prior negative BRCA testing was found to have deletion of BRCA1 exons1-2. CONCLUSIONS: The olaparib/carboplatin combination is tolerable and has modest activity in sporadic TNBC patients. Further evaluation of predictive biomarkers to identify those with BRCA wild type who had response is warranted.