Small-molecule Anaplastic Lymphoma Kinase Research Articles

Liver Toxicity Observed With Lorlatinib When Combined With Strong CYP3A Inducers: Evaluation of Cynomolgus Monkey as a Nonclinical Model for Assessing the Mechanism of Combinational Toxicity.

Lorlatinib is a potent small-molecule anaplastic lymphoma kinase inhibitor approved for the treatment of patients with nonsmall cell lung cancer. In a drug-drug interaction study in healthy human participants, liver enzyme elevations were observed when a single 100 mg dose of lorlatinib was administered after multiple doses of rifampin, a strong cytochrome P450 (CYP) 3A inducer and a pregnane X receptor (PXR) agonist. A series of in vitro and in vivo studies were conducted to evaluate potential mechanisms for the observed clinical toxicity. To investigate the involvement of CYP3A and/or PXR in the observed liver toxicity, studies were conducted in cynomolgus monkeys administered lorlatinib alone or with coadministration of multiple doses of known CYP3A inducers that are predominantly PXR agonists (rifampin, St. John's wort) or predominantly constitutive androstane receptor agonists (carbamazepine, phenytoin) and a net CYP3A inhibitory PXR agonist (ritonavir). Results from the investigative studies identified cynomolgus monkeys as a pharmacologically relevant nonclinical model, which recapitulated the elevated liver function test results observed in humans. Furthermore, liver toxicity was only observed in this model when lorlatinib was coadministered with strong CYP3A inducers, and the effects were not restricted to, or exclusively dependent upon, a PXR activation mechanism. These results generated mechanistic insights on the liver enzyme elevations observed in the clinical drug-drug interaction study and provided guidance on appropriate product safety label for lorlatinib.

Elimination of dormant, autophagic ovarian cancer cells and xenografts through enhanced sensitivity to anaplastic lymphoma kinase inhibition.

Poor outcomes for patients with ovarian cancer relate to dormant, drug-resistant cancer cells that survive after primary surgery and chemotherapy. Ovarian cancer (OvCa) cells persist in poorly vascularized scars on the peritoneal surface and depend on autophagy to survive nutrient deprivation. The authors have sought drugs that target autophagic cancer cells selectively to eliminate residual disease. By using unbiased small-interfering RNA (siRNA) screens, the authors observed that knockdown of anaplastic lymphoma kinase (ALK) reduced the survival of autophagic OvCa cells. Small-molecule ALK inhibitors were evaluated for their selective toxicity against autophagic OvCa cell lines and xenografts. Autophagy was induced by reexpression of GTP-binding protein Di-Ras3 (DIRAS3) or serum starvation and was evaluated with Western blot analysis, fluorescence imaging, and transmission electron microscopy. Signaling pathways required for crizotinib-induced apoptosis of autophagic cells were explored with flow cytometric analysis, Western blot analysis, short-hairpin RNA knockdown of autophagic proteins, and small-molecule inhibitors of STAT3 and BCL-2. Induction of autophagy by reexpression of DIRAS3 or serum starvation in multiple OvCa cell lines significantly reduced the 50% inhibitory concentration of crizotinib and other ALK inhibitors. In 2 human OvCa xenograft models, the DIRAS3-expressing tumors treated with crizotinib had significantly decreased tumor burden and long-term survival in 67% to 79% of mice. Crizotinib treatment of autophagic cancer cells further enhanced autophagy and induced autophagy-mediated apoptosis by decreasing phosphorylated STAT3 and BCL-2 signaling. Crizotinib may eliminate dormant, autophagic, drug-resistant OvCa cells that remain after conventional cytoreductive surgery and combination chemotherapy. A clinical trial of ALK inhibitors as maintenance therapy after second-look operations should be seriously considered.

505P - PD-L1 expression in ALK rearranged NSCLC: All questions answered?

BackgroundTreatment of non-small cell lung cancer (NSCLC) has undergone a paradigm shift in the last decade with the advent of targeted therapy and immunotherapy. Anaplastic Lymphoma Kinase (ALK) rearrangement and Programmed Death-Ligand 1 (PD-L1) expression are important predictive biomarkers for response to targeted therapy and immunotherapy respectively. How these two biomarkers interplay with each other is yet to be determined. Various studies have tried to establish a relationship between PD-L1 expression in tumors and treatment outcomes in ALK rearranged NSCLC, with conflicting results. There is a paucity of data regarding this relationship from the Indian subcontinent. MethodsPD-L1 expression was assessed in tissue samples of 58 patients with advanced ALK rearranged NSCLC treated between 2015 and 2018. PD-L1 expression was measured by immunohistochemistry using VENTANA PD-L1 (SP263) antibody on formalin-fixed paraffin-embedded specimens. The epidemiological profile of patients and the relationship between progression free survival (PFS) on small molecule ALK inhibitor and PD-L1 expression were noted. ResultsThe median age was 47 years (range 25-72 years) and there were 25 males (50%). Most of the patients were never smokers (88%). Thirty-seven (63.7%) of tumors expressed PD-L1 (≥1%) and 13 (22.4%) tumors had a PD-L1 expression of ≥ 50%. All of the patients received crizotinib as first line ALK inhibitor. Of the 44 patients evaluable for treatment response there was no significant difference in PFS between PDL1 negative cohort (median PFS - 13.8 months, CI 6.2-21.3 months) and PDL1 positive cohort (median PFS 10.9 months, CI 6.5-15.2 months, P-0.81). No association was seen between PD-L1 expression and PFS when PD-L1 expression levels were stratified by median or quartiles. In the subgroup analysis, PFS was not significantly different between different patterns of PDL1 expression or intensity of staining. ConclusionsIn contrast to previous studies, we found a higher incidence of PD-L1 expression in patients with ALK rearranged advanced NSCLC. PFS on ALK inhibitors did not differ based on PD-L1 expression. However, this was a retrospective study in a limited number of patients. More prospective studies are needed to confirm our findings. Legal entity responsible for the studyThe authors. FundingHas not received any funding. DisclosureAll authors have declared no conflicts of interest.

An HPLC–PDA method for determination of alectinib concentrations in the plasma of an adolescent

Alectinib is a central nervous system-active small molecule anaplastic lymphoma kinase (ALK) inhibitor that is effective in the treatment of patients with ALK positive tumors, including advanced non-small cell lung cancers and lymphomas. A simple, isocratic high-performance liquid chromatography–photo diode array detection (HPLC–PDA) assay for measurement of alectinib in human plasma is described. Alectinib is extracted from the plasma matrix by addition of methanol, followed by centrifugation and acidification with 0.1% formic acid. It elutes with a run time of 4.6 min using a 250 mm × 4.6 mm RP-C18 column with 0.1% aqueous formic acid and methanol (35:65, v/v) and a flow rate of 1 mL/min. Detection was at 339 nm. Linear calibration plots were achieved in the range of 0.1–20 μg/mL for alectinib (r2 = 0.9996). With limits of detection and quantification of 0.05 and 0.1 μg/mL, respectively, and excellent precision (%CV < 10%), accuracy (bias < ±12%), and recovery (>97%) within the 1–20 μg/mL concentration range, this assay was suitable for measuring pre-dose alectinib concentrations in an adolescent receiving 600-mg doses twice daily.

P1.13-05 Integrin β3 Inhibition Enhances the Antitumor Activity of Alk Inhibitor in Alkrearranged NSCLC

Anaplastic lymphoma kinase (ALK)-positive cancers are sensitive to small molecule ALK kinase inhibitors, but most cases experience failure following treatment. Hence, additional drug targets and combination therapeutic treatments are needed. We investigated gene expression that is regulated by the expression of ALK and explored its roles in cancer progression and therapeutic implication. We screened ALK-rearranged non-small cell lung cancer (NSCLC) cases using immunohistochemistry and fluorescence in situ hybridization, and then conducted multiplex gene expression analysis. We also performed a clinicopathological analysis to validate the findings. Additional cellular experiments, including inhibition and migration assays, and in vivo lung cancer model studies were performed. Among patients with ALK-rearranged NSCLC, integrin β3 (ITGB3) was one of the overexpressed genes in comparison with that in ALK-negative NSCLC (P = 0.0003). ALK and integrin β3 expression were positively correlated, and we discovered that high integrin β3 mRNA expression was associated with metastasis and more advanced tumor stages (P < 0.005; P < 0.05). Furthermore, we found that inhibition of both ALK and integrin β3 led to increased drug sensitivity in vitro and in vivo (both P < 0.05). We discovered a positive correlation between ALK and integrin β3 expression levels in ALK rearranged NSCLC. Our findings suggest that high integrin β3 expression in ALK-rearranged NSCLC is associated with tumor progression and a worse prognosis. This demonstrates the prognostic value of integrin β3 and provides a rationale for combination treatment with ALK and integrin β3 inhibitors in patients with ALK-rearranged NSCLC.

Integrin β3 Inhibition Enhances the Antitumor Activity of ALK Inhibitor in ALK-Rearranged NSCLC.

Purpose: Anaplastic lymphoma kinase (ALK)-positive cancers are sensitive to small-molecule ALK kinase inhibitors, but most cases experience failure following treatment. Hence, additional drug targets and combination therapeutic treatments are needed. We investigated gene expression that is regulated by the expression of ALK and explored its roles in cancer progression and therapeutic implication.Experimental Design: We screened ALK-rearranged non-small cell lung cancer (NSCLC) cases using immunohistochemistry and fluorescence in situ hybridization and then conducted multiplex gene expression analysis. We also performed a clinicopathologic analysis to validate the findings. Additional cellular experiments, including inhibition and migration assays, and in vivo lung cancer model studies were performed.Results: Among patients with ALK-rearranged NSCLC, integrin β3 (ITGB3) was one of the overexpressed genes in comparison with that in ALK-negative NSCLC (P = 0.0003). ALK and integrin β3 expression were positively correlated, and we discovered that high integrin β3 mRNA expression was associated with metastasis and more advanced tumor stages (P < 0.005; P < 0.05). Furthermore, we found that inhibition of both ALK and integrin β3 led to increased drug sensitivity in vitro and in vivo (both P < 0.05).Conclusions: We discovered a positive correlation between ALK and integrin β3 expression levels in ALK-rearranged NSCLC. Our findings suggest that high integrin β3 expression in ALK-rearranged NSCLC is associated with tumor progression and a worse prognosis. This finding demonstrates the prognostic value of integrin β3 and provides a rationale for combination treatment with ALK and integrin β3 inhibitors in patients with ALK-rearranged NSCLC. Clin Cancer Res; 24(17); 4162-74. ©2018 AACR.

SHP2 inhibition restores sensitivity in ALK-rearranged non-small-cell lung cancer resistant to ALK inhibitors.

Most anaplastic lymphoma kinase (ALK)-rearranged non-small-cell lung tumors initially respond to small-molecule ALK inhibitors, but drug resistance often develops. Of tumors that develop resistance to highly potent second-generation ALK inhibitors, approximately half harbor resistance mutations in ALK, while the other half have other mechanisms underlying resistance. Members of the latter group often have activation of at least one of several different tyrosine kinases driving resistance. Such tumors are not expected to respond to lorlatinib-a third-generation inhibitor targeting ALK that is able to overcome all clinically identified resistant mutations in ALK-and further therapeutic options are limited. Herein, we deployed a shRNA screen of 1,000 genes in multiple ALK-inhibitor-resistant patient-derived cells (PDCs) to discover those that confer sensitivity to ALK inhibition. This approach identified SHP2, a nonreceptor protein tyrosine phosphatase, as a common targetable resistance node in multiple PDCs. SHP2 provides a parallel survival input downstream of multiple tyrosine kinases that promote resistance to ALK inhibitors. Treatment with SHP099, the recently discovered small-molecule inhibitor of SHP2, in combination with the ALK tyrosine kinase inhibitor (TKI) ceritinib halted the growth of resistant PDCs through preventing compensatory RAS and ERK1 and ERK2 (ERK1/2) reactivation. These findings suggest that combined ALK and SHP2 inhibition may be a promising therapeutic strategy for resistant cancers driven by several different ALK-independent mechanisms underlying resistance.

Metabolites of alectinib in human: their identification and pharmacological activity.

Two metabolites (M4 and M1b) in plasma and four metabolites (M4, M6, M1a and M1b) in faeces were detected through the human ADME study following a single oral administration of [14C]alectinib, a small-molecule anaplastic lymphoma kinase inhibitor, to healthy subjects. In the present study, M1a and M1b, which chemical structures had not been identified prior to the human ADME study, were identified as isomers of a carboxylate metabolite oxidatively cleaved at the morpholine ring. In faeces, M4 and M1b were the main metabolites, which shows that the biotransformation to M4 and M1b represents two main metabolic pathways for alectinib. In plasma, M4 was a major metabolite and M1b was a minor metabolite. The contribution to in vivo pharmacological activity of these circulating metabolites was assessed from their in vitro pharmacological activity and plasma protein binding. M4 had a similar cancer cell growth inhibitory activity and plasma protein binding to that of alectinib, suggesting its contribution to the antitumor activity of alectinib, whereas the pharmacological activity of M1b was insignificant.

Absorption, distribution, metabolism and excretion (ADME) of the ALK inhibitor alectinib: results from an absolute bioavailability and mass balance study in healthy subjects

1. Alectinib is a highly selective, central nervous system-active small molecule anaplastic lymphoma kinase inhibitor.2. The absolute bioavailability, metabolism, excretion and pharmacokinetics of alectinib were studied in a two-period single-sequence crossover study. A 50 μg radiolabelled intravenous microdose of alectinib was co-administered with a single 600 mg oral dose of alectinib in the first period, and a single 600 mg/67 μCi oral dose of radiolabelled alectinib was administered in the second period to six healthy male subjects.3. The absolute bioavailability of alectinib was moderate at 36.9%. Geometric mean clearance was 34.5 L/h, volume of distribution was 475 L and the hepatic extraction ratio was low (0.14).4. Near-complete recovery of administered radioactivity was achieved within 168 h post-dose (98.2%) with excretion predominantly in faeces (97.8%) and negligible excretion in urine (0.456%). Alectinib and its major active metabolite, M4, were the main components in plasma, accounting for 76% of total plasma radioactivity. In faeces, 84% of dose was excreted as unchanged alectinib with metabolites M4, M1a/b and M6 contributing to 5.8%, 7.2% and 0.2% of dose, respectively.5. This novel study design characterised the full absorption, distribution, metabolism and excretion properties in each subject, providing insight into alectinib absorption and disposition in humans.

Prevalence and natural history of ALK positive non-small-cell lung cancer and the clinical impact of targeted therapy with ALK inhibitors.

Improved understanding of molecular drivers of carcinogenesis has led to significant progress in the management of lung cancer. Patients with non-small-cell lung cancer (NSCLC) with anaplastic lymphoma kinase (ALK) gene rearrangements constitute about 4%–5% of all NSCLC patients. ALK+ NSCLC cells respond well to small molecule ALK inhibitors such as crizotinib; however, resistance invariably develops after several months of treatment. There are now several newer ALK inhibitors, with the next generation of agents targeting resistance mutations. In this review, we will discuss the prevalence and clinical characteristics of ALK+ lung cancer, current treatment options, and future directions in the management of this subset of NSCLC patients.

ALK in Lung Cancer: Past, Present, and Future

In 2007, scientists discovered that anaplastic lymphoma kinase (ALK) gene rearrangements are present in a small subset of non-small-cell lung cancers. ALK-positive cancers are highly sensitive to small-molecule ALK kinase inhibitors, such as crizotinib. Phase I and II studies of crizotinib in ALK-positive lung cancer demonstrated impressive activity and clinical benefit, leading to rapid US Food and Drug Administration approval in 2011. Although crizotinib induces remissions and extends the lives of patients, cures are not achieved as resistance to therapy develops. In this review, we will discuss the history of this field, current diagnostic and treatment practices, and future challenges and opportunities to advance outcomes for patients with ALK-positive lung cancers.

Crizotinib: A comprehensive review

Anaplastic lymphoma kinase (ALK) gene rearrangements are present in a small subset of non-small-cell lung cancers. ALK-positivity confers sensitivity to small-molecule ALK kinase inhibitors, such as crizotinib. The integration of crizotinib into standard treatment practice in NSCLC will rest on the widespread implementation of an effective screening system for newly diagnosed patients with NSCLC which is flexible enough to incorporate new targets as treatments are developed for them. Phase I and II studies of crizotinib in ALK-positive lung cancer have demonstrated significant activity and impressive clinical benefit, which led to its early approval by USFDA in 2011. Although crizotinib induces remissions and extends the lives of patients, there have been reports of emerging resistance to Crizotinib therapy. In this review, we discuss the history, mechanism of action, uses, adverse effects, dose modifications and future challenges and opportunities for patients with ALK-positive lung cancers.

IS7-14 - Results of a GlobaL Phase II Study with Crizotinib in Advanced ALK-Positive Non-Small-Cell Lung Cancer (NSCLC)

ABSTRACT Background Anaplastic lymphoma kinase (ALK) is a known oncogenic driver in NSCLC, and approximately 5% of patients harbor ALK gene rearrangements. Crizotinib is a first-in-class, selective small-molecule ALK inhibitor with demonstrated clinical activity in ALK-positive NSCLC. PROFILE 1005 is a global, multicenter, open-label, single-arm phase II study evaluating the safety and efficacy of crizotinib in previously treated patients with ALK-positive advanced NSCLC. Methods Crizotinib 250 mg was administered BID to patients with ALK-positive advanced NSCLC who had received ≥1 chemotherapy for locally advanced/metastatic disease, including those with treated brain metastases. Most patients had centrally confirmed ALK-positivity by break-apart fluorescence in-situ hybridization, defined as ≥15% of tumor cells with split signals. Disease response was evaluated by RECIST 1.1 every 6 weeks. Results As of January 2012, 901 patients had been dosed. The first 261 patients who had enrolled and started crizotinib by February 2011, with a median duration of treatment of 48 weeks, were considered to be the mature population. Demographic, exposure, and efficacy results in both the overall and mature populations are provided in the table. Among all 901 patients, 15% discontinued treatment due to adverse events (AEs) and 10% had a dose reduction due to an AE. The most frequent AEs of any cause were vision disorder (54%), nausea (51%), diarrhea (44%), vomiting (44%), and constipation (37%), which were mostly grade 1/2. Conclusions Crizotinib demonstrated a high response rate that was durable, with a median progression-free survival of 8.1 months. Crizotinib has a favorable tolerability profile. These findings continue to provide strong evidence for crizotinib as a standard of care for advanced ALK-positive NSCLC. Overall population Mature population Total no. of patients, n 901 261 Response-evaluable patients, n 803 259 Demographics: Median age, years Male/female, % ECOG PS 0/1, % Adenocarcinoma, % 53 43/57 82 92 52 46/54 83 94 Duration of treatment (weeks), median (range) 20 ( 48 (1 - 94) ORR, % (95% exact CI) 46 (42–50) 60 (54–66) Duration of response (weeks), median a (95% CI) 47 (36–54) 46 (35–54) Disease control rate, % (95% exact CI): At 6 weeks At 12 weeks 80 (77–83) 61 (58–64) 86 (82–90) 75 (70–80) PFS (months), median a (95% CI) 8.1 (6.9–9.5) 8.1 (6.8–9.7) ECOG PS, Eastern Cooperative Oncology Group performance status; ORR, objective response rate; PFS, progression-free survival. a Kaplan–Meier estimate Disclosure D. Kim: Compensated advisory relationship: Pfizer. Honoraria: Pfizer. S. Lanzalone: Employment: Pfizer. Stock ownership: Pfizer. A. Polli: Employment: Pfizer. Stock ownership: Pfizer. A. Shaw: Compensated advisory relationship: Pfizer, Ariad, Chugai, Novartis, Daiichi-Sankyo. Research funding: AstraZeneca, Novartis. All other authors have declared no conflicts of interest.

440O - Results of a First-In-Human Phase I Study of the Alk Inhibitor LDK378 in Advanced Solid Tumors

ABSTRACT Background Translocations of the anaplastic lymphoma kinase (ALK) gene occur in 3–8% of NSCLC. LDK378 is a novel, potent small molecule ALK inhibitor that produces tumor regressions in ALK-driven (ALK+) NSCLC xenografts. A Phase 1 study is being conducted with the primary objective of determining the MTD and safety profile in patients (pts) with ALK+ cancers. Other objectives are to assess safety, PK, and antitumor activity in pts with ALK+ NSCLC, either previously untreated with ALK inhibitors, or relapsed following ALK inhibitor treatment, and other ALK+ cancers. Methods LDK378 was administered orally once-daily on a continuous 21-day schedule, in adult pts with advanced malignancies harboring a genetic alteration in ALK who progressed on standard therapy or for whom there was no effective therapy. Dose escalation was guided by a Bayesian logistic regression model to determine the MTD, and started at 50 mg/day. Results As of 25 April2012, 56 pts (primary site: lung 50 pts [37 with prior crizotinib]; breast 4 pts; other 2 pts; median age 53 (22–76) years; 88% ECOG PS 0/1) had received LDK378 at doses of 50–750 mg/day. Of 47 pts evaluable for response (per investigator), there were 24 (51%) responses. All responses were in ALK+ NSCLC (FISH positive in ≥15% of tumor cells). In 26 pts with NSCLC who had progressed following crizotinib and were treated at ≥400 mg/day there were 21 (81%) responses. Dose limiting toxicities (DLTs) have occurred in 2/14 pts at 400 mg/day, 2/9 pts at 600 mg/day, and 1/9 pts at 750 mg/day. DLTs included diarrhea, vomiting, nausea, dehydration, and ALT elevation. The MTD was 750 mg/day. At the cutoff date, 36 (64%) pts remain on treatment. Discontinuations were due to adverse events (AEs) in 1 (2%), and disease progression in 19 (34%) pts. The most frequent AEs (all grades) were nausea 33 (59%), vomiting 30 (54%), and diarrhea 27 (48%) pts. The most frequent Grade 3/4 AE was diarrhea (5 [9%] pts). Oral absorption of LDK378 was rapid with a Tmax of 5–6 hours, and half-life was about 36 hours. Conclusions Daily oral LDK378 is well tolerated and the MTD was 750 mg/day. Striking activity was seen in ALK+ NSCLC pts treated at doses ≥400mg, who had previously progressed following crizotinib. Disclosure A.T. Shaw: I have a consultant/advisory role for Pfizer, Novartis, Chugar, Ariad, and Daiichi. D.R. Camidge: I have a compensated consultant/advisory role with Novartis Pharmacuticals Inc. S. Sharma: I have a compensated consultant/advisory role for Novartis (LEAD Summit). I have received honoraria for Novartis LEAD Summit. I have received research funding from Novartis (Phase Ib clinical trial). D.S.W. Tan: I have received research funding from Novartis. M. Goldwasser: Employment or leadership position to disclose: Associate Director Biostatistics: Novartis Pharmaceuticals. D. Dai: Employment or leadership position to disclose. Clinical Pharmacology Expert: Novartis Pharmaceuticals; Stock ownership: Myself; Novartis Pharmaceuticals. A.L. Boral: I am an employee of Novartis Institutes for Biomedical Sciences (Executive Director). I have stock ownership (Novartis). All other authors have declared no conflicts of interest.

1268P - Visual Disturbances in Patients (PTS) with Anaplastic Lymphoma Kinase (ALK)-Positive Advanced Non-Small Cell Lung Cancer (NSCLC) Treated with Crizotinib

ABSTRACT Background Crizotinib is a potent, small-molecule ALK inhibitor with a high response rate in advanced ALK-positive NSCLC. In early phase studies, patients (pts) commonly reported mild visual disturbances. Here we characterize such visual effects with objective and subjective measures in an ongoing phase II study (PROFILE 1005). Methods Previously treated pts with advanced ALK-positive NSCLC received crizotinib 250 mg BID. Ophthalmological examinations, comprising best corrected visual acuity (BCVA), biomicroscopy, and fundoscopy, were performed at screening and after a visual effect event was reported, except for a subset of pts in France, where examinations were done every 4 cycles. In addition, on day 1 of each cycle, pts completed the Visual Symptom Assessment Questionnaire (VSAQ) that assessed the presence, frequency, timing, duration, and severity of visual effects and their impact on activities of daily living (ADL). Results As of Jan 2012, 901 pts had enrolled in PROFILE 1005; 21 - 27% were evaluable for ophthalmological examinations (visual acuity: 193/901; biomicroscopy: 239/901; fundoscopy: 239/901). There were no changes in BCVA, conjunctiva, cornea, anterior chamber, iris, lens, or fundus attributable to crizotinib. The most frequently reported ophthalmological finding was worsening of cataracts (right eye: 9%; left eye: 10%), which was not attributed to crizotinib. Visual effects as identified by the VSAQ were reported by 65% of pts (405/622) at cycle 2 (C2), 58% at C3 (323/558), 55% at C4 (287/519), and 50% at C5 (247/495). The most common were appearance of flashing lights, streamers/strings/floaters, and overlapping shadows. Most pts reported each event as lasting ≤1 min (C2: 63%; C3: 68%; C4: 72%; C5: 75%), occurring mostly in the morning (46–59%) and/or evening (70–74%). The majority of pts reported that visual effects were not at all or a little bothersome, with no or minimal impact on ADL. Similar results were observed in the French pt subset. Conclusions There were no objective ophthalmological changes associated with the visual effects on crizotinib. Pt-reported visual effects were frequent, but transient, with no or minimal impact on ADL. Disclosure B. Besse: Research funding: Pfizer. B. Solomon: Compensated advisory relationship: Pfizer. Research funding: Pfizer. A. Shaw: Compensated advisory relationship: Pfizer, Ariad, Chugai, Novartis, and Daiichi-Sankyo. Research funding: AstraZeneca and Novartis. D. Kim: Compensated advisory relationship: Pfizer. Honoraria: Pfizer. R. Schachar: Compensated employment: Pfizer. Stock ownership: Pfizer. K. Wilner: Compensated employment: Pfizer. Stock ownership: Pfizer. A. Reisman: Compensated employment: Pfizer. Stock ownership: Pfizer. C.H. Bartlett: Compensated employment: Pfizer. Stock ownership: Pfizer. S. Iyer: Compensated employment: Pfizer. All other authors have declared no conflicts of interest.

1230PD - Updated Results of a Global Phase II Study with Crizotinib in Advanced Alk-Positive Non-Small Cell Lung Cancer (NSCLC)

ABSTRACT Background Anaplastic lymphoma kinase (ALK) is a known oncogenic driver in NSCLC, and approximately 5% of patients harbor ALK gene rearrangements. Crizotinib is a first-in-class, selective small-molecule ALK inhibitor with demonstrated clinical activity in ALK-positive NSCLC. PROFILE 1005 is a global, multicenter, open-label, single-arm phase II study evaluating the safety and efficacy of crizotinib in previously treated patients with ALK-positive advanced NSCLC. Methods Crizotinib 250 mg was administered BID to patients with ALK-positive advanced NSCLC who had received ≥1 chemotherapy for locally advanced/metastatic disease, including those with treated brain metastases. Most patients had centrally confirmed ALK-positivity by break-apart fluorescence in-situ hybridization, defined as ≥15% of tumor cells with split signals. Disease response was evaluated by RECIST 1.1 every 6 weeks. Results As of January 2012, 901 patients had been dosed. The first 261 patients who had enrolled and started crizotinib by February 2011, with a median duration of treatment of 48 weeks, were considered to be the mature population. Demographic, exposure, and efficacy results in both the overall and mature populations are provided in the table. Among all 901 patients, 15% discontinued treatment due to adverse events (AEs) and 10% had a dose reduction due to an AE. The most frequent AEs of any cause were vision disorder (54%), nausea (51%), diarrhea (44%), vomiting (44%), and constipation (37%), which were mostly grade 1/2. Conclusions Crizotinib demonstrated a high response rate that was durable, with a median progression-free survival of 8.1 months. Crizotinib has a favorable tolerability profile. These findings continue to provide strong evidence for crizotinib as a standard of care for advanced ALK-positive NSCLC. Overall population Mature population Total no. of patients, n 901 261 Response-evaluable patients, n 803 259 Demographics: Median age, years Male/female, % ECOG PS 0/1, % Adenocarcinoma, % 53 43/57 82 92 52 46/54 83 94 Duration of treatment (weeks), median (range) 20 ( 48 (1 - 94) ORR, % (95% exact CI) 46 (42–50) 60 (54–66) Duration of response (weeks), median a (95% CI) 47 (36–54) 46 (35–54) Disease control rate, % (95% exact CI): At 6 weeks At 12 weeks 80 (77–83) 61 (58–64) 86 (82–90) 75 (70–80) PFS (months), median a (95% CI) 8.1 (6.9–9.5) 8.1 (6.8–9.7) ECOG PS, Eastern Cooperative Oncology Group performance status; ORR, objective response rate; PFS, progression-free survival. a Kaplan–Meier estimate Disclosure D. Kim: Compensated advisory relationship: Pfizer. Honoraria: Pfizer. S. Lanzalone: Employment: Pfizer. Stock ownership: Pfizer. A. Polli: Employment: Pfizer. Stock ownership: Pfizer. A. Shaw: Compensated advisory relationship: Pfizer, Ariad, Chugai, Novartis, Daiichi-Sankyo. Research funding: AstraZeneca, Novartis. All other authors have declared no conflicts of interest.

The R1275Q Neuroblastoma Mutant and Certain ATP-competitive Inhibitors Stabilize Alternative Activation Loop Conformations of Anaplastic Lymphoma Kinase

Anaplastic lymphoma kinase (ALK) is a receptor tyrosine kinase that, when genetically altered by mutation, amplification, chromosomal translocation or inversion, has been shown to play an oncogenic role in certain cancers. Small molecule inhibitors targeting the kinase activity of ALK have proven to be effective therapies in certain ALK-driven malignancies and one such inhibitor, crizotinib, is now approved for the treatment of EML4-ALK-driven, non-small cell lung cancer. In neuroblastoma, activating point mutations in the ALK kinase domain can drive disease progression, with the two most common mutations being F1174L and R1275Q. We report here crystal structures of the ALK kinase domain containing the F1174L and R1275Q mutations. Also included are crystal structures of ALK in complex with novel small molecule ALK inhibitors, including a classic type II inhibitor, that stabilize previously unobserved conformations of the ALK activation loop. Collectively, these structures illustrate a different series of activation loop conformations than has been observed in previous ALK crystal structures and provide insight into the activating nature of the R1275Q mutation. The novel active site topologies presented here may also aid the structure-based drug design of a new generation of ALK inhibitors.

First-in-human phase I study of the ALK inhibitor LDK378 in advanced solid tumors.

3007 Background: LDK378 is a novel, potent and selective small molecule anaplastic lymphoma kinase (ALK) inhibitor (IC50 0.00015 μM), that does not inhibit c-MET (IC50 3.2 μM). Tumor regression has been observed in ALK-driven NSCLC xenografts. A first-in-human, Phase I study is being conducted to determine the MTD and safety profile in patients (pts) with tumors with ALK rearrangement, amplification or mutation. Other objectives were safety, PK and antitumor activity in pts with ALK-driven NSCLC, both naïve to ALK inhibitors and relapsed following previous ALK inhibitor treatment, and other ALK-positive cancers. Methods: Adult pts with advanced malignancies harboring a genetic alteration in ALK who progressed on standard therapy or for whom there was no effective therapy, were given once daily oral LDK378 on a continuous 21-day schedule. Dose escalation, starting at 50 mg/day, was guided by a Bayesian logistic regression model (BLRM) to determine the MTD. Results: At a January 5th 2012 cutoff,31 pts (primary site: lung 26 pts; breast 3 pts; other 2 pts; median age 52 years; 87% ECOG PS 0/1) were enrolled and received LDK378 at doses of 50–750 mg/day. Two dose limiting toxicities, Grade (Gr) 3 alanine aminotransferase elevation (1 pt), and Gr 3 hypophosphatemia (1 pt) occurred in 8 pts at a 400 mg dose level. BLRM allowed dose escalation, and there were no DLTs in 4 pts at 500 mg. Median duration of treatment with LDK378 was 7 weeks (range &lt;1–22+). At the cutoff date, 13 (42%) pts discontinued treatment: 1 (3%) due to adverse events (AEs), and 12 (39%) due to disease progression; 18 (58%) pts were still on treatment. The most frequent AEs (all Gr) were nausea (45%), vomiting (36%), and diarrhea (29%). The most frequent Gr 3/4 AEs were diarrhea and dyspnea (2 pts [7%] each). At doses ≥400 mg steady state exposures exceeded efficacious exposures in xenograft models. Of 16 pts with available response data (RECIST, per investigator) there were 4/6 responses in crizotinib (CRZ)-treated pts, and 2/10 responses in CRZ-naïve pts, of whom 7 were treated below 400 mg. All responses were in NSCLC. Conclusions: Daily oral LDK378 is well tolerated up to 500 mg/day, and escalation continues at 750 mg/day. Preliminary responses have been seen in both CRZ-naïve and CRZ-relapsed pts.

Development of treatment strategies for advanced neuroblastoma

Neuroblastoma is the most common cancer in childhood. The majority of patients with neuroblastoma are assigned to the high-risk group based on age at diagnosis, stage, histology, MYCN status, and DNA ploidy. Their prognosis remains unsatisfactory; the 5-year event-free survival (EFS) rate is generally 40 %. During the past 20 years, much effort has been made to reinforce chemotherapy, including the introduction of high-dose chemotherapy with autologous stem cell rescue, resulting in a 5-year EFS rate of around 30 %. Subsequently, maintenance therapy aimed at eradicating residual tumors after induction and consolidation therapies was introduced, consisting of differentiation-inducing agents, retinoids, and immunotherapy using anti-GD2 antibodies combined with cytokines. However, such additional treatment provided benefit to only 10-20 % of patients, while the prognosis of about half the patients remains poor. Currently, novel targeted agents are under development. Among them, anaplastic lymphoma kinase (ALK) inhibitors and aurora kinase A inhibitors are promising. ALK somatic mutation or gene amplification predisposing neuroblastoma development occurs in up to 15 % of neuroblastomas. Crizotinib is a dual-specific inhibitor of ALK/Met and inhibits proliferation of neuroblastoma cells harboring R1275Q-mutated ALK or amplified wild-type ALK, but not cells harboring F1174L. Instead, cells with F1174L are sensitive to another small molecule ALK inhibitor, TAE684. Aurora kinase A plays a pivotal role in centrosome maturation and spindle formation during mitosis. MLN8237 (alisertib) is a small molecule inhibitor of aurora kinase A that is currently in early-phase clinical testing. Future treatment will be individually planned, adapting targeted agents based on personal biological tumor characteristics.

Mechanisms of Resistance to Crizotinib in Patients with ALK Gene Rearranged Non–Small Cell Lung Cancer

Patients with anaplastic lymphoma kinase (ALK) gene rearrangements often manifest dramatic responses to crizotinib, a small-molecule ALK inhibitor. Unfortunately, not every patient responds and acquired drug resistance inevitably develops in those who do respond. This study aimed to define molecular mechanisms of resistance to crizotinib in patients with ALK(+) non-small cell lung cancer (NSCLC). We analyzed tissue obtained from 14 patients with ALK(+) NSCLC showing evidence of radiologic progression while on crizotinib to define mechanisms of intrinsic and acquired resistance to crizotinib. Eleven patients had material evaluable for molecular analysis. Four patients (36%) developed secondary mutations in the tyrosine kinase domain of ALK. A novel mutation in the ALK domain, encoding a G1269A amino acid substitution that confers resistance to crizotinib in vitro, was identified in two of these cases. Two patients, one with a resistance mutation, exhibited new onset ALK copy number gain (CNG). One patient showed outgrowth of epidermal growth factor receptor (EGFR) mutant NSCLC without evidence of a persistent ALK gene rearrangement. Two patients exhibited a KRAS mutation, one of which occurred without evidence of a persisting ALK gene rearrangement. One patient showed the emergence of an ALK gene fusion-negative tumor compared with the baseline sample but with no identifiable alternate driver. Two patients retained ALK positivity with no identifiable resistance mechanism. Crizotinib resistance in ALK(+) NSCLC occurs through somatic kinase domain mutations, ALK gene fusion CNG, and emergence of separate oncogenic drivers.