Yakov Rotshteyn

Antiviral activity of single-dose PRO 140, a CCR5 monoclonal antibody, in HIV-infected adults

BACKGROUND The current goal of human immunodeficiency virus type 1 (HIV-1) therapy is to maximally suppress viral replication. Securing this goal requires new drugs and treatment classes. The chemokine receptor CCR5 provides an entry portal for HIV-1, and PRO 140 is a humanized monoclonal antibody that binds to CCR5 and potently inhibits CCR5-tropic (R5) HIV-1 in vitro. METHODS A randomized, double-blind, placebo-controlled, dose-escalating study was conducted in 39 individuals with HIV-1 RNA levels or =5000 copies/mL, CD4(+) cell counts > or =250 cells/microL, no antiretroviral therapy for 3 months, and only R5 HIV-1 detectable. Cohorts were randomized 3:10 to receive placebo or doses of PRO 140 of 0.5, 2, or 5 mg/kg. Subjects were monitored for 58 days for safety, antiviral effects, and serum concentrations of PRO 140. RESULTS PRO 140 was generally well tolerated and demonstrated potent, rapid, prolonged, and dose-dependent antiviral activity. Mean reductions in HIV-1 RNA level of 0.58 log(10), 1.20 log(10) (P= .0002) and 1.83 log(10) (P= .0001) were observed for the 0.5-, 2-, and 5-mg/kg dose groups, respectively. Reductions in mean viral load of > or =10-fold were observed within 4 days and persisted for 2-3 weeks after treatment. CONCLUSIONS This trial established clear proof of concept for PRO 140 as a potent antiretroviral agent with extended activity after a single dose. TRIAL REGISTRATION ISRCTN Register: ISRCTN45537485 .

Phase 2a Study of the CCR5 Monoclonal Antibody PRO 140 Administered Intravenously to HIV-Infected Adults

ABSTRACT The anti-CCR5 antibody PRO 140 has shown potent and prolonged antiretroviral activity in subjects infected with CCR5-tropic (R5) HIV-1. Prior studies have examined single intravenous doses ranging up to 5 mg/kg of body weight or up to three subcutaneous doses ranging up to 324 mg. Here we report the results of a randomized, double-blind, placebo-controlled trial that examined the antiviral activity, tolerability, and pharmacokinetics of single 5-mg/kg and 10-mg/kg intravenous infusions of PRO 140 in 31 treated subjects. Eligibility criteria included HIV-1 RNA levels of >5,000 copies/ml, CD4+ cell counts of >300/μl, no antiretroviral therapy for ≥12 weeks, and detection of only R5 HIV-1 in the original Trofile assay. Following poststudy testing with an enhanced-sensitivity Trofile assay, one subject treated with 10 mg/kg was reclassified as having dual/mixed-tropic virus at screening, and the data for that subject were censored from efficacy analyses. The mean maximum reduction of the HIV-1 RNA level from the baseline level was 1.8 log10 units for both the 5-mg/kg and 10-mg/kg doses (P < 0.0001 relative to placebo). Viral loads reached their nadir at day 12 posttreatment and remained significantly (P < 0.01) reduced through day 29 for both PRO 140 dose groups. Treatment was generally well tolerated, with no dose-limiting toxicity being observed. Peak serum concentrations and overall exposures increased proportionally with dose. In summary, single 5-mg/kg and 10-mg/kg doses of PRO 140 exhibited potent, long-lived antiviral activity and were generally well tolerated. The findings further delineate the safety and antiviral properties of this novel, long-acting antiretroviral agent.

Anti-HIV-1 activity of weekly or biweekly treatment with subcutaneous PRO 140, a CCR5 monoclonal antibody.

BACKGROUND PRO 140 is a humanized CCR5 monoclonal antibody that has demonstrated potent antiviral activity when it is administered intravenously to adults infected with CCR5-tropic (R5) human immunodeficiency virus type 1 (HIV-1). This study is the first to evaluate subcutaneous administration. METHODS A randomized, double-blind, placebo-controlled study was conducted among 44 subjects with HIV-1 RNA levels of >5000 copies/mL, CD4(+) cell counts of >300 cells/microL, no receipt of antiretroviral therapy for >or=12 weeks, and only R5 HIV-1 detectable. Subjects received placebo, 162 mg of PRO 140, or 324 mg of PRO 140 weekly for 3 weeks or 324 mg of PRO 140 every other week for 2 doses by means of subcutaneous infusion. Subjects were monitored for 58 days for safety, antiviral effects, and PRO 140 serum concentrations. RESULTS Subcutaneous PRO 140 demonstrated potent and prolonged antiretroviral activity. Mean log(10) reductions in HIV-1 RNA level were 0.23, 0.99 (P=.009), 1.37 (P<.001), and 1.65 (P<.001) for the placebo, 162 mg weekly, 324 mg biweekly, and 324 mg weekly dose groups, respectively. Viral loads remained suppressed between successive doses. Treatment was generally well tolerated. CONCLUSIONS This trial demonstrates proof of concept for a monoclonal antibody administered subcutaneously in HIV-1 infected individuals. Subcutaneous PRO 140 offers the potential for significant dose-dependent HIV-1 RNA suppression and infrequent patient self-administration. TRIAL REGISTRATION ClinicalTrials.gov identifier: NCT00642707 .

Novel Small-Molecule Inhibitors of Hepatitis C Virus Entry Block Viral Spread and Promote Viral Clearance in Cell Culture

Combinations of direct-acting anti-virals offer the potential to improve the efficacy, tolerability and duration of the current treatment regimen for hepatitis C virus (HCV) infection. Viral entry represents a distinct therapeutic target that has been validated clinically for a number of pathogenic viruses. To discover novel inhibitors of HCV entry, we conducted a high throughput screen of a proprietary small-molecule compound library using HCV pseudoviral particle (HCVpp) technology. We independently discovered and optimized a series of 1,3,5-triazine compounds that are potent, selective and non-cytotoxic inhibitors of HCV entry. Representative compounds fully suppress both cell-free virus and cell-to-cell spread of HCV in vitro. We demonstrate, for the first time, that long term treatment of an HCV cell culture with a potent entry inhibitor promotes sustained viral clearance in vitro. We have confirmed that a single amino acid variant, V719G, in the transmembrane domain of E2 is sufficient to confer resistance to multiple compounds from the triazine series. Resistance studies were extended by evaluating both the fusogenic properties and growth kinetics of drug-induced and natural amino acid variants in the HCVpp and HCV cell culture assays. Our results indicate that amino acid variations at position 719 incur a significant fitness penalty. Introduction of I719 into a genotype 1b envelope sequence did not affect HCV entry; however, the overall level of HCV replication was reduced compared to the parental genotype 1b/2a HCV strain. Consistent with these findings, I719 represents a significant fraction of the naturally occurring genotype 1b sequences. Importantly, I719, the most relevant natural polymorphism, did not significantly alter the susceptibility of HCV to the triazine compounds. The preclinical properties of these triazine compounds support further investigation of entry inhibitors as a potential novel therapy for HCV infection.

Abstract B215: Correlation of PSMA ADC exposure with reduction in tumor growth rate determined using serial PSA measurements from a Phase I clinical trial.

Background: PSMA ADC is a fully humanized antibody to PSMA conjugated to the potent microtubule disrupting agent monomethyl auristatin E (MMAE). The safety and PK of PSMA ADC as well as changes in PSA levels were evaluated in a Phase I dose escalation study in patients with taxane-refractory metastatic castration-resistant prostate cancer (mCRPC) (PSMA ADC 1301). Stein et al. (2010) has previously demonstrated correlation between growth rate derived from serial PSA measurements with survival in patients with mCRPC and suggested that growth rate could serve as a novel endpoint indicative of response to therapy and overall survival. Methods: The PK parameters for PSMA ADC, Total antibody (Ab) and free MMAE obtained from PSMA ADC Study 1301 (N=52) were calculated using non-compartmental analysis. The final data set contains 253 longitudinal PSA observations from 52 subjects dosed with 0.4-2.8 mg/kg PSMA ADC. The Stein model was re-expressed as a 7-parameter population model to describe the longitudinal PSA data which includes: Baseline PSA level (Y0), PSA decrease rate (D), PSA increase rate (G), inter-patient variability for Y0, G and D respectively, and residual variance. Model parameters were estimated using Phoenix/NLME®. Exposure-efficacy relationships and correlation between G and covariates were determined. Results: PSMA ADC, Total Ab, and MMAE exposure increased linearly with increasing PSMA ADC dose. Longitudinal PSA data were correctly fitted to the re-expressed Stein model. G continuously decreased with increasing PSMA ADC exposure, suggesting an exposure dependent drug effect. There was no apparent saturation of this effect at the maximum observed exposure, and no influence of age, baseline albumin and ALT/AST levels on G was evident. Conclusions: In a Phase I study where PSMA ADC (0.4-2.8 mg/kg) was administered to patients with mCRPC, we observed a correlation of PSMA ADC exposure with the reduction of G, suggesting that PSMA ADC may provide dose-dependent therapeutic benefit to subjects with mCRPC. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B215. Citation Format: Yakov Rotshteyn, F Mercier, R Bruno, Nancy Stambler, Robert J. Israel, Vivien Wong. Correlation of PSMA ADC exposure with reduction in tumor growth rate determined using serial PSA measurements from a Phase I clinical trial. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B215.

Abstract 4492: Targeting Ras-mutated tumors with novel multiplex PI3K inhibitors

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Background: The simultaneous dysregulation of both PI3K and Ras-MAPK pathways is characteristic of some of the most aggressive forms of human cancer (e.g. tumors with Ras mutations). It has been demonstrated in preclinical studies that effective treatment of Ras-mutated tumors requires blockade of both pathways. Therefore, the ability to achieve this outcome with a single agent holds great clinical promise. One key function of the PI3K and Ras-MAPK pathways is the regulation of protein translation. Both pathways converge to regulate eIF-4E, a key factor in cap-dependent translation of mRNAs encoding critical proteins involved in tumorigenesis and tumor cell survival (e.g. c-Myc and Mcl-1). The PI3K pathway activates eIF-4E via mTOR-mediated phosphorylation and suppression of 4E-BP1, a negative regulator of eIF-4E; the Ras-MAPK pathway modulates eIF4E function through phosphorylation by MNK. It has recently been demonstrated that MNK expression and MNK-dependent phosphorylation of eIF-4E on Ser209 are required for Ras-induced oncogenic transformation, and for tumor development and progression in vivo. Using a combination of rational drug design and conditional lethal screening, we discovered a novel lead series of small molecule inhibitors with multiplex activities against PI3K, mTOR and MNK. Aspects of the preclinical characterization of these novel multiplex PI3K inhibitors will be presented. Methods and Results: Multiplex PI3K inhibitors are potent inhibitors of tumor cell proliferation and this anti-proliferative activity is independent of the presence of oncogenic mutations (e.g. Ras, PI3K, B-Raf or EGFR). Multiplex PI3K inhibitors effectively induced cell killing of tumor cell lines, including those with Ras mutations, at concentrations ranging from 100 nM to single digit μM, and are superior to either PI3K or MEK inhibitors used alone or in combination. Mechanistically, multiplex PI3K inhibitors inhibit the phosphorylation of AKT, 4E-BP1, ribosomal S6 and eIF-4E, inhibit the expression of c-Myc and Mcl-1 proteins, and induce caspase activity. In human tumor xenograft efficacy studies, multiplex PI3K inhibitors demonstrated robust in vivo anti-tumor activity. The levels of key pharmacodynamic endpoints (phosphorylated AKT, 4E-BP1, ribosomal S6 and eIF-4E) were significantly reduced in treated tumors, consistent with the proposed mechanism of action. Conclusion: Newly-discovered novel multiplex PI3K inhibitors are capable of simultaneously targeting both PI3K and Ras-MAPK pathways and exhibit potent anti-tumor activity in Ras-mutated tumors. These inhibitors warrant further investigation for development as a potentially novel class of anti-cancer drugs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4492. doi:10.1158/1538-7445.AM2011-4492