Activity of eftozanermin alfa plus venetoclax in preclinical models and patients with acute myeloid leukemia.

Activation of apoptosis in malignant cells is an established strategy for controlling cancer and is potentially curative. To assess the impact of concurrently inducing the extrinsic and intrinsic apoptosis-signaling pathways in acute myeloid leukemia (AML), we evaluated activity of the TRAIL receptor agonistic fusion protein eftozanermin alfa (eftoza; ABBV-621) in combination with the B-cell lymphoma protein-2 selective inhibitor venetoclax in preclinical models and human patients. Simultaneously stimulating intrinsic and extrinsic apoptosis-signaling pathways with venetoclax and eftoza, respectively, enhanced their activities in AML cell lines and patient-derived ex vivo/in vivo models. Eftoza activity alone or plus venetoclax required death receptor 4/5 (DR4/DR5) expression on the plasma membrane but was independent of TP53 or FLT3-ITD status. The safety/tolerability of eftoza as monotherapy and in combination with venetoclax was demonstrated in patients with relapsed/refractory AML in a phase 1 clinical trial. Treatment-related adverse events were reported in 2 of 4 (50%) patients treated with eftoza monotherapy and 18 of 23 (78%) treated with eftoza plus venetoclax. An overall response rate of 30% (7/23; 4 complete responses [CRs], 2 CRs with incomplete hematologic recovery, and 1 morphologic leukemia-free state) was reported in patients who received treatment with eftoza plus venetoclax and 67% (4/6) in patients with myoblasts positive for DR4/DR5 expression; no tumor responses were observed with eftoza monotherapy. These data indicate that combination therapy with eftoza plus venetoclax to simultaneously activate the extrinsic and intrinsic apoptosis-signaling pathways may improve clinical benefit compared with venetoclax monotherapy in relapsed/refractory AML with an acceptable toxicity profile. This trial was registered at www.clinicaltrials.gov as #NCT03082209.

Overall survival with sacituzumab govitecan in hormone receptor-positive and human epidermal growth factor receptor 2-negative metastatic breast cancer (TROPiCS-02): a randomised, open-label, multicentre, phase 3 trial.

BACKGROUND: Sacituzumab govitecan demonstrated significant progression-free survival benefit over chemotherapy in the phase 3 TROPiCS-02 trial in patients with pretreated, endocrine-resistant hormone receptor-positive, human epidermal growth factor receptor 2-negative (HR+ and HER2-) metastatic breast cancer with limited treatment options. Here, we report the protocol-specified final analysis of overall survival and endpoints by trophoblast cell-surface antigen 2 (Trop-2) expression and other variables. METHODS: In this randomised, open-label, multicentre, phase 3 trial, which took place in 91 centres across North America (the USA and Canada) and Europe (Belgium, France, Germany, Italy, the Netherlands, Spain, and the UK), patients were randomly assigned (1:1) to receive sacituzumab govitecan or chemotherapy (eribulin, vinorelbine, capecitabine, or gemcitabine). Patients had confirmed HR+ and HER2- locally recurrent inoperable or metastatic breast cancer and had received at least one previous endocrine therapy, a taxane, and a CDK4/6 inhibitor in any setting and two to four previous chemotherapy regimens for metastatic disease. The primary endpoint was progression-free survival (previously reported and not included in this analysis), and secondary endpoints included overall survival, objective response rate (ORR), and patient-reported outcomes. Overall survival was assessed using stratified log-rank tests and Cox regression. Trop-2 expression was assessed in tumour tissue by immunohistochemistry. In the statistical testing hierarchy, ORR and patient-reported outcomes were tested sequentially if overall survival was significant. This study is registered with ClinicalTrials.gov, NCT03901339. FINDINGS: At the data cutoff date of July 1, 2022, 543 of 776 screened patients were randomly assigned between May 30, 2019, and April 5, 2021, with 272 patients in the sacituzumab govitecan group and 271 patients in the chemotherapy group. With a 12·5-month (IQR 6·4-18·8) median follow-up, 390 deaths occurred among 543 patients. Overall survival was significantly improved with sacituzumab govitecan versus chemotherapy (median 14·4 months [95% CI 13·0-15·7] vs 11·2 months [10·1-12·7]; hazard ratio [HR] 0·79, 95% CI 0·65-0·96; p=0·020); survival benefit was consistent across Trop-2 expression-level subgroups. ORR was significantly improved with sacituzumab govitecan compared with chemotherapy (57 [21%] patients vs 38 [14%]; odds ratio 1·63 [95% CI 1·03-2·56]; p=0·035), as was time to deterioration of global health status and quality of life (median 4·3 months vs 3·0 months; HR 0·75 [0·61-0·92]; p=0·0059) and fatigue (median 2·2 months vs 1·4 months; HR 0·73 [0·60-0·89]; p=0·0021). The safety profile of sacituzumab govitecan was consistent with previous studies (including the TROPiCS-02 primary analysis and the ASCENT trial). One fatal adverse event (septic shock caused by neutropenic colitis) was determined to be related to sacituzumab govitecan treatment. INTERPRETATION: Sacituzumab govitecan demonstrated statistically significant and clinically meaningful benefit over chemotherapy, with a 3·2-month median overall survival improvement and a manageable safety profile. These data support sacituzumab govitecan as a new treatment option for patients with pretreated, endocrine-resistant HR+ and HER2- metastatic breast cancer. FUNDING: Gilead Sciences.

Eftozanermin alfa (ABBV-621) monotherapy in patients with previously treated solid tumors: findings of a phase 1, first-in-human study.

Eftozanermin alfa (eftoza), a second-generation tumor necrosis factor-related apoptosis-inducing ligand receptor (TRAIL-R) agonist, induces apoptosis in tumor cells by activation of death receptors 4/5. This phase 1 dose-escalation/dose-optimization study evaluated the safety, pharmacokinetics, pharmacodynamics, and preliminary activity of eftoza in patients with advanced solid tumors. Patients received eftoza 2.5-15 mg/kg intravenously on day 1 or day 1/day 8 every 21 days in the dose-escalation phase, and 1.25-7.5 mg/kg once-weekly (QW) in the dose-optimization phase. Dose-limiting toxicities (DLTs) were evaluated during the first treatment cycle to determine the maximum tolerated dose (MTD) and recommended phase 2 dose (RP2D). Pharmacodynamic effects were evaluated in circulation and tumor tissue. A total of 105 patients were enrolled in the study (dose-escalation cohort, n = 57; dose-optimization cohort, n = 48 patients [n = 24, colorectal cancer (CRC); n = 24, pancreatic cancer (PaCA)]). In the dose-escalation cohort, seven patients experienced DLTs. MTD and RP2D were not determined. Most common treatment-related adverse events were increased alanine aminotransferase and aspartate aminotransferase levels, nausea, and fatigue. The one treatment-related death occurred due to respiratory failure. In the dose-optimization cohort, three patients (CRC, n = 2; PaCA, n = 1) had a partial response. Target engagement with regard to receptor saturation, and downstream apoptotic pathway activation in circulation and tumor were observed. Eftoza had acceptable safety, evidence of pharmacodynamic effects, and preliminary anticancer activity. The 7.5-mg/kg QW regimen was selected for future studies on the basis of safety findings, pharmacodynamic effects, and biomarker modulations. (Trial registration number: NCT03082209 (registered: March 17, 2017)).

Phase Ib Study of Telisotuzumab Vedotin in Combination With Erlotinib in Patients With c-Met Protein-Expressing Non-Small-Cell Lung Cancer.

PURPOSE: Overexpression of c-Met protein and epidermal growth factor receptor (EGFR) mutations can co-occur in non-small-cell lung cancer (NSCLC), providing strong rationale for dual targeting. Telisotuzumab vedotin (Teliso-V), a first-in-class antibody-drug conjugate targeting c-Met, has shown a tolerable safety profile and antitumor activity as monotherapy. Herein, we report the results of a phase Ib study (ClinicalTrials.gov identifier: NCT02099058) evaluating Teliso-V plus erlotinib, an EGFR tyrosine kinase inhibitor (TKI), in patients with c-Met-positive (+) NSCLC. PATIENTS AND METHODS: This study evaluated Teliso-V (2.7 mg/kg once every 21 days) plus erlotinib (150 mg once daily) in adult patients (age ≥ 18 years) with c-Met+ NSCLC. Later enrollment required presence of an EGFR-activating mutation (EGFR-M+) and progression on a prior EGFR TKI. End points included safety, pharmacokinetics, objective response rate (ORR), and progression-free survival (PFS). The efficacy-evaluable population consisted of c-Met+ patients (confirmed histology [H]-score ≥ 150) who had at least one postbaseline scan; c-Met+ patients with H-scores ≥ 225 were classified as c-Met high. RESULTS: As of January 2020, 42 patients were enrolled (N = 36 efficacy-evaluable). Neuropathies were the most common any-grade adverse events reported, with 24 of 42 patients (57%) experiencing at least one event. The pharmacokinetic profile of Teliso-V plus erlotinib was similar to Teliso-V monotherapy. Median PFS for all efficacy-evaluable patients was 5.9 months (95% CI, 2.8 to not reached). ORR for EGFR-M+ patients (n = 28) was 32.1%. Of EGFR-M+ patients, those who were c-Met high (n = 15) had an ORR of 52.6%. Median PFS was 6.8 months for non-T790M+ and for those whose T790M status was unknown, versus 3.7 months for T790M+. CONCLUSION: Teliso-V plus erlotinib showed encouraging antitumor activity and acceptable toxicity in EGFR TKI-pretreated patients with EGFR-M+, c-Met+ NSCLC.

A Phase 1b Study of Telisotuzumab Vedotin in Combination With Nivolumab in Patients With NSCLC.

INTRODUCTION: Telisotuzumab vedotin (Teliso-V) is an anti-c-Met-directed antibody-drug conjugate that has exhibited antitumor activity as monotherapy in NSCLC. Its potential activity combined with programmed cell death protein-1 inhibitors has not been previously evaluated. METHODS: In a phase 1b study (NCT02099058), adult patients (≥18 y) with advanced NSCLC received combination therapy with Teliso-V (1.6, 1.9, or 2.2 mg/kg, every 2 wk) plus nivolumab (3 mg/kg, 240 mg, or per locally approved label). The primary objective was to assess safety and tolerability; secondary objectives included the evaluation of antitumor activity. RESULTS: As of January 2020, a total of 37 patients received treatment with Teliso-V (safety population) in combination with nivolumab; 27 patients (efficacy population) were c-Met immunohistochemistry-positive. Programmed death-ligand 1 (PD-L1) status was evaluated in the efficacy population (PD-L1-positive [PD-L1+]: n = 15; PD-L1-negative [PD-L1-]: n = 9; PD-L1-unknown: n = 3). The median age was 67 years and 74% (20 of 27) of patients were naive to immune checkpoint inhibitors. The most common any-grade treatment-related adverse events were fatigue (27%) and peripheral sensory neuropathy (19%). The pharmacokinetic profile of Teliso-V plus nivolumab was similar to Teliso-V monotherapy. The objective response rate was 7.4%, with two patients (PD-L1+, c-Met immunohistochemistry H-score 190, n = 1; PD-L1-, c-Met H-score 290, n = 1) having a confirmed partial response. Overall median progression-free survival was 7.2 months (PD-L1+: 7.2 mo; PD-L1-: 4.5 mo; PD-L1-unknown: not reached). CONCLUSIONS: Combination therapy with Teliso-V plus nivolumab was well tolerated in patients with c-Met+ NSCLC with limited antitumor activity.

Co-clinical Modeling of the Activity of the BET Inhibitor Mivebresib (ABBV-075) in AML.

BACKGROUND/AIM: The therapeutic potential of bromodomain and extra-terminal motif (BET) inhibitors in hematological cancers has been well established in preclinical and early-stage clinical trials, although as of yet, no BETtargeting agent has achieved approval. To add insight into potential response to mivebresib (ABBV-075), a broadspectrum BET inhibitor, co-clinical modeling of individual patient biopsies was conducted in the context of a Phase I trial in acute myeloid leukemia (AML). MATERIALS AND METHODS: Co-clinical modeling involves taking the patient's biopsy and implanting it in mice with limited passage so that it closely retains the original characteristics of the malignancy and allows comparisons of response between animal model and clinical data. Procedures were developed, initially with neonate NOD/Shi-scid-IL2rγnull (NOG) mice and then optimized with juvenile NOG-EXL as host mice, eventually resulting in a robust rate of engraftment (16 out of 26, 62%). RESULTS: Results from the co-clinical AML patient-derived xenograft (PDX) modeling (6 with >60% inhibition of bone marrow blasts) were consistent with the equivalent clinical data from patients receiving mivebresib in monotherapy, and in combination with venetoclax. The modeling system also demonstrated the activity of a novel BD2-selective BET inhibitor (ABBV-744) in the preclinical AML setting. Both agents were also highly effective in inhibiting blast counts in the spleen (10/10 and 5/6 models, respectively). CONCLUSION: These findings confirm the validity of the model system in the co-clinical setting, establish highly relevant in vivo models for the discovery of cancer therapy, and indicate the therapeutic value of BET inhibitors for AML and, potentially, myelofibrosis treatment.

Phase I Open-Label Study Evaluating the Safety, Pharmacokinetics, and Preliminary Efficacy of Dilpacimab in Patients with Advanced Solid Tumors.

Dilpacimab (formerly ABT-165), a novel dual-variable domain immunoglobulin, targets both delta-like ligand 4 (DLL4) and VEGF pathways. Here, we present safety, pharmacokinetic (PK), pharmacodynamic (PD), and preliminary efficacy data from a phase I study (trial registration ID: NCT01946074) of dilpacimab in patients with advanced solid tumors. Eligible patients (≥18 years) received dilpacimab intravenously on days 1 and 15 in 28-day cycles at escalating dose levels (range, 1.25-7.5 mg/kg) until progressive disease or unacceptable toxicity. As of August 2018, 55 patients with solid tumors were enrolled in the dilpacimab monotherapy dose-escalation and dose-expansion cohorts. The most common treatment-related adverse events (TRAE) included hypertension (60.0%), headache (30.9%), and fatigue (21.8%). A TRAE of special interest was gastrointestinal perforation, occurring in 2 patients (3.6%; 1 with ovarian and 1 with prostate cancer) and resulting in 1 death. The PK of dilpacimab showed a half-life ranging from 4.9 to 9.5 days, and biomarker analysis demonstrated that the drug bound to both VEGF and DLL4 targets. The recommended phase II dose for dilpacimab monotherapy was established as 3.75 mg/kg, primarily on the basis of tolerability through multiple cycles. A partial response was achieved in 10.9% of patients (including 4 of 16 patients with ovarian cancer). The remaining patients had either stable disease (52.7%), progressive disease (23.6%), or were deemed unevaluable (12.7%). These results demonstrate that dilpacimab monotherapy has an acceptable safety profile, with clinical activity observed in patients with advanced solid tumors.

Phase I Study of 2- or 3-Week Dosing of Telisotuzumab Vedotin, an Antibody-Drug Conjugate Targeting c-Met, Monotherapy in Patients with Advanced Non-Small Cell Lung Carcinoma.

PURPOSE: Telisotuzumab vedotin (Teliso-V) is an anti-c-Met-directed antibody-drug conjugate. Here, we present safety and efficacy data from a phase I/Ib study of Teliso-V monotherapy evaluated in once every 2 weeks/once every 3 weeks schedules in patients with non-small cell lung cancer (NSCLC). PATIENTS AND METHODS: During dose escalation, patients received Teliso-V monotherapy intravenously once every 3 weeks (0.15-3.3 mg/kg) or once every 2 weeks (1.6-2.2 mg/kg). The dose-expansion phase enrolled patients with NSCLC and c-Met H-score ≥150 (c-Met+) or MET amplification/exon 14 skipping mutations. Safety, pharmacokinetics, and efficacy were assessed. Herein, the analysis of patients receiving ≥1.6 mg/kg once every 2 weeks or ≥2.4 mg/kg once every 3 weeks Teliso-V is reported. RESULTS: Fifty-two patients with NSCLC were enrolled and received ≥1.6 mg/kg Teliso-V once every 2 weeks (n = 28) or ≥2.4 mg/kg Teliso-V once every 3 weeks (n = 24). The most common adverse events were fatigue (54%), peripheral neuropathy (42%), and nausea (38%). No dose-limiting toxicities were observed for Teliso-V once every 2 weeks and once every 3 weeks up to 2.2 and 2.7 mg/kg, respectively. The recommended phase II dose was established at 1.9 mg/kg once every 2 weeks and 2.7 mg/kg once every 3 weeks on the basis of overall safety and pharmacokinetics. Forty of 52 patients were c-Met+ (33 nonsquamous, 6 squamous, 1 mixed histology) and were included in the efficacy-evaluable population. Of those, 9 (23%) had objective responses with median duration of response of 8.7 months; median progression-free survival was 5.2 months. CONCLUSIONS: Teliso-V monotherapy was tolerated and showed antitumor activity in c-Met+ NSCLC. On the basis of overall safety, pharmacokinetics, and efficacy outcomes, 1.9 mg/kg Teliso-V once every 2 weeks and 2.7 mg/kg once every 3 weeks schedules were selected for further clinical development.

Phase I Dose-Escalation and -Expansion Study of Telisotuzumab (ABT-700), an Anti-c-Met Antibody, in Patients with Advanced Solid Tumors.

This first-in-human phase I study evaluated the pharmacokinetics, safety, and preliminary efficacy of telisotuzumab, formerly called ABT-700, an antagonistic antibody directed against c-Met. For dose escalation (3+3 design), 3 to 6 patients with advanced solid tumors were enrolled into four dose cohorts (5-25 mg/kg). In the dose-expansion phase, a subset of patients was prospectively selected for MET amplification (FISH screening). Patients received telisotuzumab intravenously on day 1 every 21 days. For dose expansion, 15 mg/kg was chosen as the dose on the basis of safety, pharmacokinetics, and other data from the escalation cohorts. Forty-five patients were enrolled and received at least one dose of telisotuzumab (dose escalation, n = 15; dose expansion, n = 30). Telisotuzumab showed a linear pharmacokinetics profile; peak plasma concentration was proportional to dose level. There were no acute infusion reactions and no dose-limiting toxicities were observed. The most common treatment-related adverse events included hypoalbuminemia (n = 9, 20.0%) and fatigue (n = 5, 11.1%). By Response Evaluation Criteria In Solid Tumors (RECIST), 4 of 10 (40.0%) patients with MET-amplified tumors had confirmed partial response in target lesions (one ovarian, two gastric, and one esophageal), two (20.0%) had stable disease, three (30.0%) had progressive disease; one patient was unable to be evaluated. Among patients with nonamplified tumors (n = 35), no objective responses were observed; however, 11 patients had stable disease per RECIST criteria. In conclusion, telisotuzumab has an acceptable safety profile with clinical activity observed in patients with MET-amplified advanced solid tumors.

Flavopiridol targets c-KIT transcription and induces apoptosis in gastrointestinal stromal tumor cells.

Gastrointestinal stromal tumors (GIST) are characterized by activating mutations in the c-KIT gene which confers ligand-independent activation of the KIT receptor. Imatinib mesylate has been shown to effectively block constitutively active KIT and delay tumor growth. However, resistance to imatinib mesylate is emerging as a major clinical problem and novel therapies are needed. We report that treatment of GIST cells with the transcriptional inhibitor flavopiridol, initially down-regulates the antiapoptotic proteins bcl-2, mcl-1, and X-linked inhibitor of apoptosis protein which occurs as early as 4 hours after exposure. This is followed at 24 hours by the transcriptional suppression of KIT resulting in poly(ADP-ribose) polymerase cleavage and apoptosis. To separate the apoptotic effect of KIT suppression relative to the down-regulation of antiapoptotic proteins, we used small interfering RNA-directed knockdown of KIT. Results show that focused suppression of KIT alone is sufficient to induce apoptosis in GIST cells, but not to the same extent as flavopiridol. In contrast, imatinib mesylate, which inhibits KIT kinase activity but does not suppress total KIT expression, fails to cause apoptosis. We also show that flavopiridol suppresses KIT mRNA expression through positive transcriptional elongation factor inhibition and decreases KIT promoter activity. This causes a global decrease in the level of functionally mature KIT at the cell surface, resulting in a decrease in autophosphorylation at tyrosine residues 703 and 721, which characterizes activated KIT. Our results indicate that targeting KIT expression and these antiapoptotic proteins with flavopiridol represents a novel means to disrupt GIST cell dependence on KIT signaling and collectively renders these cells sensitive to apoptosis.

First-in-Human Phase I, Dose-Escalation and -Expansion Study of Telisotuzumab Vedotin, an Antibody-Drug Conjugate Targeting c-Met, in Patients With Advanced Solid Tumors.

PURPOSE: This first-in-human study evaluated telisotuzumab vedotin (Teliso-V), formerly called ABBV-399, an antibody-drug conjugate of the anti-c-Met monoclonal antibody ABT-700 and monomethyl auristatin E. MATERIALS AND METHODS: For dose escalation, three to six patients with advanced solid tumors were enrolled in eight cohorts (0.15 to 3.3 mg/kg). The dose-expansion phase enrolled patients with non-small-cell lung cancer (NSCLC) with c-Met-overexpressing tumors (c-Met positive; immunohistochemistry membrane H-score ≥ 150). Patients received Teliso-V monotherapy intravenously on day 1 once every 3 weeks. Safety, tolerability, pharmacokinetics, and maximum tolerated dose were determined. RESULTS: Forty-eight patients were enrolled (median age, 65 years; 35.4% NSCLC; median four prior therapies). One patient each in the 3.0-mg/kg (n = 9) and 3.3-mg/kg (n = 3) cohorts experienced dose-limiting toxicities. Although the maximum tolerated dose was not formally identified, the recommended phase II dose was defined as 2.7 mg/kg on the basis of overall safety and tolerability. The most frequent treatment-emergent adverse events (any grade) were fatigue (42%), nausea (27%), constipation (27%), decreased appetite (23%), vomiting (21%), dyspnea (21%), diarrhea (19%), peripheral edema (19%), and neuropathy (17%). The most frequent Teliso-V-related grade ≥ 3 adverse events were fatigue, anemia, neutropenia, and hypoalbuminemia (4% each). Teliso-V and total antibody pharmacokinetics were approximately dose proportional, with a mean harmonic half-life of 2 to 4 days each. Prospective screening identified 35 (60%) of 58 patients with c-Met-positive NSCLC. Of 16 patients with c-Met-positive NSCLC who were treated with Teliso-V 2.4 to 3.0 mg/kg, three (18.8%; 95% CI, 4.1% to 45.7%) achieved a partial response (median response duration, 4.8 months; median progression-free survival, 5.7 months; 95% CI, 1.2 months to 15.4 months). No other patients experienced a response. CONCLUSION: Teliso-V monotherapy demonstrated favorable safety and tolerability profiles, with encouraging evidence of antitumor activity in patients with c-Met-positive NSCLC.

A First-Time-in-Human Study of GSK2636771, a Phosphoinositide 3 Kinase Beta-Selective Inhibitor, in Patients with Advanced Solid Tumors.

Background: The PI3K/protein kinase B (AKT) pathway is commonly activated in several tumor types. Selective targeting of p110ß could result in successful pathway inhibition while avoiding the on- and off-target effects of pan-PI3K inhibitors. GSK2636771 is a potent, orally bioavailable, adenosine triphosphate-competitive, selective inhibitor of PI3Kß.Methods: We evaluated the safety, pharmacokinetics, pharmacodynamics and antitumor activity of GSK2636771 to define the recommended phase II dose (RP2D). During the dose-selection and dose-escalation stages (parts 1 and 2), patients with PTEN-deficient advanced solid tumors received escalating doses of GSK2636771 (25-500 mg once daily) using a modified 3+3 design to determine the RP2D; tumor type-specific expansion cohorts (part 3) were implemented to further assess tumor responses at the RP2D.Results: A total of 65 patients were enrolled; dose-limiting toxicities were hypophosphatemia and hypocalcemia. Adverse events included diarrhea (48%), nausea (40%), and vomiting (31%). Single- and repeat-dose exposure increased generally dose proportionally. GSK2636771 400 mg once daily was the RP2D. Phospho/total AKT ratio decreased with GSK2636771 in tumor and surrogate tissue. A castrate-resistant prostate cancer (CRPC) patient harboring PIK3CB amplification had a partial response for over a year; an additional 10 patients derived durable (≥24 weeks) clinical benefit, including two other patients with CRPC with PIK3CB alterations (≥34 weeks). GSK2636771 400 mg once daily orally induced sufficient exposure and target inhibition with a manageable safety profile.Conclusions: Genomic aberrations of PIK3CB may be associated with clinical benefit from GSK2636771. Clin Cancer Res; 23(19); 5981-92. ©2017 AACR.

Drg1 expression in 131 colorectal liver metastases: correlation with clinical variables and patient outcomes.

PURPOSE: The differentiation-related gene-1 (Drg1) is a recently identified gene down-regulated in malignancy and a putative suppressor of colorectal cancer metastases. Its expression is associated with improved survival in patients with prostate or breast cancer. Drg1 expression is also associated with resistance to irinotecan therapy in preclinical colorectal cancer models. The clinical evaluation of Drg1 in colorectal cancer has been limited. We performed this study to evaluate the role of Drg1 in a large cohort of patients with metastatic colorectal cancer who were irinotecan naive. EXPERIMENTAL DESIGN: We examined Drg1 expression by immunohistochemistry in 131 patients with metastatic colorectal cancer enrolled in a clinical trial of adjuvant fluorouracil-based therapy from 1991 to 1995. We correlated expression of Drg1 to numerous clinical and tumor related variables and to patient outcomes, including a subset of patients who recurred and received irinotecan-based therapy. RESULTS: Drg1 expression was identified in all metastatic tissue samples. There was a trend for unilobar metastases with high Drg1 expression (P = 0.07) and a suggestion of improved 2-year survival (82.4% versus 69.6%, P = 0.148). High Drg1 expression suggested irinotecan resistance (P = 0.07). CONCLUSIONS: In colorectal cancer, Drg1 expression may be associated with a less aggressive, indolent colorectal cancer. High Drg1 may also be associated with relative resistance to irinotecan. The role of Drg1 in malignancy continues to be defined.

A phase I clinical trial of the sequential combination of irinotecan followed by flavopiridol.

PURPOSE: Flavopiridol potently enhances the effect of irinotecan with cures in colorectal cancer xenografts, and is associated with modulation of several molecular targets, including p21, Differentiation-related gene 1 (Drg1), and p53. We initiated a phase I trial of the sequential combination of irinotecan followed by flavopiridol to determine the maximal tolerated dose of this combination therapy. PATIENTS AND METHODS: Forty-five patients with advanced solid tumors were enrolled. Irinotecan was administered first (100 or 125 mg/m(2)) followed 7 hours later by escalating flavopiridol (10-70 mg/m(2)) given weekly over 1 hour for 4 of 6 weeks. At the maximal tolerated dose, the pharmacokinetic analysis was expanded and pre- and posttreatment tumor biopsies were done. RESULTS: At irinotecan 100 mg/m(2), dose-limiting diarrhea and myelosuppression were observed with flavopiridol 70 mg/m(2). At irinotecan 125 mg/m(2), we observed dose-limiting hyperbilirubinemia, fatigue, and myelosuppression at flavopiridol 60 mg/m(2). Peak flavopiridol concentrations of >/=2 mumol/L were achieved above flavopiridol 50 mg/m(2). No significant pharmacokinetic interactions with irinotecan were noted. Baseline serum bilirubin significantly predicted cycle 1 dose-limiting toxicity and neutropenia. We observed partial responses in three patients and prolonged stable disease (i.e., >6 months) in 36% of patients including adrenocortical cancer and hepatocellular cancer. Patients with wild-type p53 and either no change or low posttreatment biopsy p21 and a decrease in Drg1 expression showed stable or responsive disease to the combination therapy. CONCLUSIONS: The recommended phase II dose with irinotecan 100 mg/m(2) is flavopiridol 60 mg/m(2) and with irinotecan 125 mg/m(2) is flavopiridol 50 mg/m(2). Toxicity can be predicted by baseline bilirubin. Clinical activity is encouraging and may correlate to changes in p21 and Drg1 levels in patients with wild type p53 tumors following therapy.

Drg1, a novel target for modulating sensitivity to CPT-11 in colon cancer cells.

Treatment of the human colon cancer cells Hct116 with SN-38 (an active metabolite of CPT-11) resulted in G2 cell cycle arrest without induction of apoptosis. However, subsequent treatment of SN-38-treated Hct116 cells with flavopiridol induced apoptosis. One of the genes markedly up-regulated during cell cycle arrest by SN-38 and suppressed during apoptosis by SN-38 followed by flavopiridol in Hct116 cells is Drg1. We found that Drg1 had profound effects on SN-38 sensitivity. Inhibition of endogenous Drg1 expression in Hct116 cells by stable expression of an antisense (AS) Drg1 cDNA increased the sensitivity of cells to undergo apoptosis by SN-38. Clonogenic and apoptosis assays with AS Drg1-expressing cells showed a 2-fold decrease in the IC50 and a 4-5-fold increase in induction of apoptosis with SN-38. Conversely, the forced expression of Drg1 in SW620 cells increased the resistance of these cells to SN-38-induced apoptosis by 2-5-fold. Moreover, when xenografted in mice, AS Drg1-expressing Hct116 cells were 3-fold more sensitive to CPT-11 as compared with vector transfected Hct116 cells. Similarly, tumors established from Drg1 overexpressing SW620 cells were more resistant to CPT-11 as compared with tumors established from vector-transfected SW620 cells in mice. Taken together, our data suggest that Drg1 is a novel gene that plays a direct role in resistance to CPT-11. Inhibition of Drg1 may provide a new means to increase the sensitivity of colon cancer cells to CPT-11.

The cyclin-dependent kinase inhibitor flavopiridol potentiates gamma-irradiation-induced apoptosis in colon and gastric cancer cells.

PURPOSE: Flavopiridol is a cyclin-dependent kinase inhibitor currently under development by the National Cancer Institute both as a single agent and in combination with chemotherapy. There have been numerous reports that flavopiridol potently enhances the induction of apoptosis by chemotherapy. However, the effect of flavopiridol on radiotherapy (RT)-induced apoptosis has been largely untested. RT has become the cornerstone of adjuvant treatment of colorectal and gastric cancer. In view of this, we elected to evaluate the effect of flavopiridol on potentiating RT-induced apoptosis in the human colon cancer cell line HCT-116 and the gastric cancer cell line MKN-74. EXPERIMENTAL DESIGN: The efficacy of combination of gamma-irradiation and flavopiridol was tested in vitro in MKN-74 and HCT-116 cells and correlated to changes in p21 expression. HCT-116 cells were also established as tumors in nude mice and treated with gamma-irradiation and flavopiridol either as single agents or in sequential combinations such that flavopiridol was either given 7 h before, concomitantly, or 3 and 7 h after gamma-irradiation. RESULTS: Flavopiridol significantly enhanced the induction of apoptosis by gamma-irradiation in both cell lines as measured by quantitative fluorescent microscopy, caspase-3 activation, poly(ADP-ribose) polymerase cleavage, and cytochrome c release. To achieve the best effect, it was important to expose the tumor cells to gamma-irradiation before the flavopiridol. This sequence dependence was confirmed in vivo. When gamma-irradiation was administered 7 h before flavopiridol, 42% of the tumor-bearing animals were rendered disease free, compared with no animals treated with either gamma-irradiation or flavopiridol alone. Examination of the p21 status of HCT-116 and MKN-74 cells, after treatment with sequential gamma-irradiation and flavopiridol, indicated a loss of p21 protein expression. Loss of p21 was mainly due to cleavage by caspases. HCT-116 cells that lack p21 (p21(-/-)) also exhibited sensitization to gamma-irradiation and showed an even greater enhancement of gamma-irradiation-induced apoptosis by flavopiridol when compared with the parental HCT-116 cells. CONCLUSIONS: These studies indicate that gamma-irradiation followed by flavopiridol enhances apoptosis and yields significantly increased tumor regressions and cures that are not achievable with radiation alone. These results indicate that flavopiridol can potently enhance the effect of gamma-radiation both in vitro and in vivo and may provide a new means to treat patients with locally advanced gastrointestinal cancers.

Flavopiridol enhances the effect of docetaxel in vitro and in vivo in human gastric cancer cells.

Gastric cancer is one of the leading causes of cancer death throughout the world. It is a disease in desperate need of new therapeutic approaches. Docetaxel, a semisynthetic taxane, has shown potent activity against a broad range of solid tumors. However, in gastric cancer, response rates to docetaxel remain only approximately 20%. In these studies we show that flavopiridol, a cyclin-dependent kinase inhibitor, potentiates docetaxel-induced apoptosis 3-fold in MKN-74 human gastric cells. This effect is sequence dependent, such that flavopiridol must follow docetaxel to induce this effect. Docetaxel induces transient arrest in the M phase of the cell cycle. Cells exit mitosis in a specific time window without cytokinesis with a decrease in cyclin B1/cdc-2 kinase activity and MPM-2 labeling. Flavopiridol treatment of docetaxel-treated cells enhances the exit from mitosis with a more rapid decrease in mitotic markers including MPM-2 labeling and cyclin B1/cdc2 kinase activity. In contrast, pretreatment with flavopiridol prevents cells from entering mitosis by inhibiting cyclin B1/cdc-2 kinase activity, thus antagonizing the docetaxel effect. The testing of this combination against MKN-74 xenografts confirms the sequence dependency. Treatment of MKN-74 tumor-bearing xenografts with docetaxel at a dose of 10 mg/kg followed 3-7 h later by flavopiridol at a dose of 2.5 mg/kg resulted in a 1-18% decrease in tumor volume. In contrast, treatment with docetaxel alone at this same dose resulted in a 394% increase in tumor volume. When flavopiridol was given immediately after docetaxel, the effect was not statistically different from that of docetaxel alone. The reverse combination of flavopiridol followed 7 h later by docetaxel was similar to treatment with docetaxel alone. Flavopiridol alone had no effect in this tumor model. Thus, flavopiridol, when combined with docetaxel in a sequence-specific manner, may provide a completely new therapeutic approach in the treatment of gastric cancer.

Phase I study of the MEK inhibitor trametinib in combination with the AKT inhibitor afuresertib in patients with solid tumors and multiple myeloma.

PURPOSE: To identify the maximum tolerated dose (MTD) and recommended Phase II dose of MEK/AKT inhibitor combination of trametinib and afuresertib. PATIENTS AND METHODS: Eligibility criteria were advanced solid tumors, 18 years or older, Eastern Cooperative Oncology Group performance status 0 or 1, and adequate organ function. Exclusion criteria included Type 1 diabetes, active GI disease, leptomeningeal disease, or current evidence/risk of retinal venous occlusion/central serous retinopathy. Clinical safety parameters and response were evaluated and analyzed. RESULTS: Twenty patients were enrolled. Dose-limiting toxicities (Grade 2 esophagitis; Grade 3 aspartate aminotransferase increased, mucosal inflammation and hypokalemia) were reported at starting dose (1.5 mg trametinib/50 mg afuresertib once daily continuously), exceeding the MTD. Subsequent de-escalation cohorts (1.5 mg/25 mg or 1.0 mg/50 mg trametinib/afuresertib) were defined as MTDs for continuous dosing. Intermittent dosing schedule [1.5 mg trametinib (continuous)/50 mg afuresertib (Days 1-10 every 28 days)] was evaluated and considered tolerable. No patients were enrolled in Phase II. The most common adverse events reported (≥10 % of all patients) included: diarrhea (60 %), dermatitis acneiform (55 %), maculo-papular rash (45 %), fatigue (30 %), dry skin (25 %), nausea (25 %), dyspnea (20 %), and vomiting (20 %). One partial response (BRAF wild-type melanoma) was reported; four patients had stable disease as best response. CONCLUSION: Continuous daily dosing of trametinib/afuresertib combination was poorly tolerated. Evaluation of intermittent dose schedule showed greater tolerability. Given the interest in combination treatment regimens of MAPK and PI3K/AKT pathway inhibitors, further study of intermittent dose schedule or combination of trametinib with more selective inhibitors may be warranted.

Combined BRAF and MEK Inhibition With Dabrafenib and Trametinib in BRAF V600-Mutant Colorectal Cancer.

PURPOSE: To evaluate dabrafenib, a selective BRAF inhibitor, combined with trametinib, a selective MEK inhibitor, in patients with BRAF V600-mutant metastatic colorectal cancer (mCRC). PATIENTS AND METHODS: A total of 43 patients with BRAF V600-mutant mCRC were treated with dabrafenib (150 mg twice daily) plus trametinib (2 mg daily), 17 of whom were enrolled onto a pharmacodynamic cohort undergoing mandatory biopsies before and during treatment. Archival tissues were analyzed for microsatellite instability, PTEN status, and 487-gene sequencing. Patient-derived xenografts were established from core biopsy samples. RESULTS: Of 43 patients, five (12%) achieved a partial response or better, including one (2%) complete response, with duration of response > 36 months; 24 patients (56%) achieved stable disease as best confirmed response. Ten patients (23%) remained in the study > 6 months. All nine evaluable during-treatment biopsies had reduced levels of phosphorylated ERK relative to pretreatment biopsies (average decrease ± standard deviation, 47% ± 24%). Mutational analysis revealed that the patient achieving a complete response and two of three evaluable patients achieving a partial response had PIK3CA mutations. Neither PTEN loss nor microsatellite instability correlated with efficacy. Responses to dabrafenib plus trametinib were comparable in patient-derived xenograft-bearing mice and the biopsied lesions from each corresponding patient. CONCLUSION: The combination of dabrafenib plus trametinib has activity in a subset of patients with BRAF V600-mutant mCRC. Mitogen-activated protein kinase signaling was inhibited in all patients evaluated, but to a lesser degree than observed in BRAF-mutant melanoma with dabrafenib alone. PIK3CA mutations were identified in responding patients and thus do not preclude response to this regimen. Additional studies targeting the mitogen-activated protein kinase pathway in this disease are warranted.

Lack of mannose-binding lectin-A enhances survival in a mouse model of acute septic peritonitis.

The mannose-binding lectin (MBL) (also known as the mannose-binding protein) is a serum protein that plays a role as an "ante-antibody" in innate immunity. In man, MBL is encoded by a single gene, whereas in mice there are two homologous proteins, MBL-A and MBL-C. In order to evaluate the relative roles of these two forms of MBL, we created MBL-A null mice that were MBL-C sufficient. We found MBL-A null mice had enhanced survival in a septic peritonitis model compared to wild-type mice and complement 3 null mice at 24 h, 48 h and 10 d (P < 0.05). Reconstitution of these mice with human MBL reversed the phenotype. Surviving mice had significantly decreased TNF-alpha and IL-6 levels in the blood and peritoneal cavity (P < 0.01). In vitro studies indicate that bacteria opsonized with MBL-A-deficient serum induced significantly less cytokine by peritoneal macrophages compared to those with wild-type serum. Our results indicate that MBL-A is a modulator of inflammation in vivo and in vitro in the mouse and that the role of MBL may extend beyond its role as an opsonin.