Jeremy Force

Blood-Based Biomarkers of SU11248 Activity and Clinical Outcome in Patients with Metastatic Imatinib-Resistant Gastrointestinal Stromal Tumor

Purpose: There is an unmet need for noninvasive markers to measure the biological effects of targeted agents, particularly those inhibiting the vascular endothelial growth factor (VEGF) receptor (VEGFR) pathway, and identify patients most likely to benefit from treatment. In this study, we investigated potential blood-based biomarkers for SU11248 (sunitinib malate), a multitargeted tyrosine kinase inhibitor, in patients with metastatic imatinib-refractory gastrointestinal stromal tumors. Experimental Design: Patients (n = 73) enrolled in a phase I/II trial received SU11248 daily for 14 or 28 days followed by 14 days without treatment per cycle. Clinical benefit was defined as progression-free survival of >6 months. We assessed plasma markers, including VEGF and soluble VEGFR-2 (sVEGFR-2), and two cellular populations bearing VEGF receptors: monocytes and, in a subset of patients, mature circulating endothelial cells (CEC). Results: Compared to patients with progressive disease, patients with clinical benefit had significantly greater increases in CECs (0.52 versus −−0.01 CEC/μL/d, P = 0.03) and smaller decreases in monocyte levels (47% versus 60%, P = 0.007) during cycle 1. VEGF increased by 2.2-fold and sVEGFR-2 decreased 25% during the first 2 weeks of treatment. Neither plasma marker correlated with clinical outcome although a modest inverse correlation was observed between sVEGFR-2 changes and plasma drug levels. Monocytes, VEGF, and sVEGFR-2 all rebounded towards baseline off treatment. Conclusions: Monocytes, VEGF, and sVEGFR-2 were consistently modulated by treatment, suggesting that they may serve as pharmacodynamic markers for SU11248. Changes in CECs and monocytes, but not the plasma markers, differed between the patients with clinical benefit and those with progressive disease. These end points merit further investigation in future trials to determine their utility as markers of SU11248 activity and clinical benefit in gastrointestinal stromal tumors and other tumor types.

Differential effects of vascular endothelial growth factor receptor-2 inhibitor ZD6474 on circulating endothelial progenitors and mature circulating endothelial cells: Implications for use as a surrogate marker of antiangiogenic activity

Purpose: Circulating endothelial cells (CEC) comprise at least two distinct populations: bone marrow–derived circulating endothelial progenitors (CEP) and mature CECs derived from existing vasculature. We hypothesized that antiangiogenic agents may have differential effects on CEPs and mature CECs and that these changes may serve as a marker of biological activity. Experimental Design: The effect of angiogenesis inhibitors on CECs was evaluated by flow cytometry after vascular endothelial growth factor (VEGF)–induced mobilization and in mice bearing Lewis lung carcinoma (LLC). Tumor angiogenesis was evaluated in parallel by immunohistochemistry. Results: In nontumor-bearing mice, VEGF administration increased both mature CECs and CEPs. This increase was inhibited by the VEGF receptor 2 inhibitor ZD6474 as well as the VEGF inhibitor–soluble Flt-1. ZD6474 had no significant effect on CECs in the absence of exogenous VEGF stimulation. In contrast, LLC-bearing mice had an increase in mature CECs but not CEPs after 3 days of treatment with ZD6474. The increase in mature CECs was dose-dependent, accompanied by a decrease in tumor microvessel density, and preceded reduction in tumor volume. Treatment of LLC-bearing mice with the vascular targeting agent ZD6126 also increased mature CECs. Conclusions: VEGF inhibitors can have differential effects on mature CECs and CEPs, and agents inhibiting tumor angiogenesis may cause a concomitant increase in mature CECs. This increase occurs in tumor-bearing but not in nontumor-bearing mice, suggesting that tumor endothelium is a potential source of mature CECs. Therefore, assessing both mature CECs and CEPs may provide insights into the mechanism of antiangiogenic agents and serve as an early surrogate marker of biological activity.

Therapeutic efficacy of endostatin exhibits a biphasic dose-response curve.

We show here that recombinant endostatin protein has a biphasic effect on the inhibition of endothelial cell migration in vitro. In tumor-bearing animals, there is a similar biphasic effect on the inhibition of tumor growth and on circulating endothelial cells after once-daily s.c. injections. This biphasic effect is revealed as a U-shaped curve in which efficacy is optimal between very low and very high doses depending on the tumor type. This result may be applicable to other inhibitors of endothelial growth and to angiogenesis. Furthermore, these results have important implications for clinicians who administer angiogenesis inhibitors for cancer or other angiogenesis-dependent diseases. When these results are taken together with two previous reports of angiogenesis inhibitors with a U-shaped dose-response, they suggest that other regulators of endothelial growth may display a similar pattern.

Evaluation of KRAS mutations, angiogenic biomarkers and DCE-MRI in patients with advanced non-small cell lung cancer receiving sorafenib

Purpose: Sorafenib, a multikinase inhibitor targeting Raf and VEGFR, has shown activity in unselected patients with non–small-cell lung cancer (NSCLC). At present there are no validated biomarkers indicative of sorafenib activity. Experimental Design: Patients received sorafenib 400 mg BID daily to determine activity and tolerability and to measure its biological effects. KRAS mutation status (N = 34), angiogenesis markers (VEGF, bFGF, FLT-1, PLGF-1) and imaging with DCE-MRI (dynamic contrast enhanced MRI) to determine early changes in tumor vascular characteristics were evaluated. Three parameters Ktrans, Kep, and Ve were measured by DCE-MRI at baseline and day 14 of cycle 1. Cytokine analysis was done on days 0, 14, 28, and 54. Results: Thirty-seven patients with previously treated stage IV NSCLC were enrolled in this single-center phase II trial. In 34 evaluable patients, 2 had partial responses and 20 had stable disease for 3 to 17 months, a disease control rate of 65%. The median progression-free survival (PFS) was 3.4 months, and median overall survival (OS) was 11.6 months. Toxicity was consistent with the known side effects of sorafenib. KRAS (32%) and EGFR mutations (22%) showed no correlation with response, PFS, or OS. Kep, was significant in predicting an improvement in OS (P = 0.035) and PFS (P = 0.029). Cytokine analysis demonstrated an improved OS for bFGF day 0 (<6 vs. >6 pg/mL; P = 0.042), whereas a PFS benefit was seen with bFGF at day 28 (<6 vs. >6; P = 0.028). Conclusions: KRAS and EGFR mutational status showed no correlation with response, PFS, or OS. Radiologic and cytokine changes may act as biomarkers indicative of early angiogenesis inhibition. Clin Cancer Res; 17(5); 1190–9. ©2011 AACR.

Regression of Drug-Resistant Lung Cancer by the Combination of Rosiglitazone and Carboplatin

Purpose: Current therapy for lung cancer involves multimodality therapies. However, many patients are either refractory to therapy or develop drug resistance. KRAS and epidermal growth factor receptor (EGFR) mutations represent some of the most common mutations in lung cancer, and many studies have shown the importance of these mutations in both carcinogenesis and chemoresistance. Genetically engineered murine models of mutant EGFR and KRAS have been developed that more accurately recapitulate human lung cancer. Recently, using cell-based experiments, we showed that platinum-based drugs and the antidiabetic drug rosiglitazone (PPARγ ligand) interact synergistically to reduce cancer cell and tumor growth. Here, we directly determined the efficacy of the PPARγ/carboplatin combination in these more relevant models of drug resistant non–small cell lung cancer. Experimental Design: Tumorigenesis was induced by activation of either mutant KRAS or EGFR. Mice then received either rosiglitazone or carboplatin monotherapy, or a combination of both drugs. Change in tumor burden, pathology, and evidence of apoptosis and cell growth were assessed. Results: Tumor burden remained unchanged or increased in the mice after monotherapy with either rosiglitazone or carboplatin. In striking contrast, we observed significant tumor shrinkage in mice treated with these drugs in combination. Immunohistochemical analyses showed that this synergy was mediated via both increased apoptosis and decreased proliferation. Importantly, this synergy between carboplatin and rosiglitazone did not increase systemic toxicity. Conclusions: These data show that the PPARγ ligand/carboplatin combination is a new therapy worthy of clinical investigation in lung cancers, including those cancers that show primary resistance to platinum therapy or acquired resistance to targeted therapy.

Nodular Regenerative Hyperplasia After Treatment With Trastuzumab Emtansine

Introduction Trastuzumab emtansine (T-DM1) is a novel antibody-drug conjugate consisting of three components: trastuzumab, a recombinant antiepidermal growth factor receptor 2 (HER2) monoclonal antibody; thioether linker; and mertansine (DM1), a maytansinoid. The trastuzumab moiety of this antibody-drug conjugate binds to HER2 on tumor cell surfaces and on internalization, the DM1 moiety is released and binds to tubulin, thereby disrupting microtubule assembly/disassembly dynamics, resulting in cell-cycle arrest and cell death. Pharmacokinetic studies reveal complete proteolysis of the polypeptide backbone of the antibody component to its constituent amino acids, resulting in lysine-linker-DM1 inside the target cell. This major metabolite was reported to have low cytotoxic potency toward hepatocytes because of inability to penetrate the hepatocyte via the charged groups of the lysine residue. Preclinical studies demonstrated generation of lysine-linker-DM1 in liver tissue, mild increase in liver enzymes, and minimal degeneration of hepatocytes in monkeys. Furthermore, the liver was believed to be immune to cytotoxic effects of tubulin-binding antimitotic agents, such as DM1, because the hepatocytes are fully differentiated and nondividing. This drug is currently undergoing evaluation through several phase II and phase III studies in patients with HER2-positive breast cancer. Recently, a large phase III clinical trial (An Open-Label Study of Trastuzumab Emtansine [T-DM1] Versus Capecitabine Lapatinib in Patients With HER2Positive Locally Advanced or Metastatic Breast Cancer [EMILIA]) reported an improvement in progression-free and overall survival with T-DM1 when compared with the combination of lapatinib and capecitabine for patients who had experienced disease progression while receiving previous trastuzumab-based therapy. In this study, the T-DM1 arm had an incidence of AST elevation of 22.4%, ALT elevation of 16.9%, and thrombocytopenia of 28%, resulting in dose reduction or discontinuation. Here we report two heavily pretreated patients with metastatic breast cancer who received T-DM1–based regimens while participating in phase Ib/II clinical trials and developed noncirrhotic portal hypertension (NCPH) as a result of nodular regenerative hyperplasia (NRH).

Assessment of objective responses using volumetric evaluation in advanced thymic malignancies and metastatic non-small cell lung cancer.

Introduction: Measurement of tumor response by standard response criteria is challenging in thymic malignancies, especially when the pleura is involved, as it often is in stage IV disease. In this study, we aimed to determine the effectiveness of volumetric response evaluation criteria in solid tumors (volumetrics) for evaluating response in patients with thymic malignancies treated on a phase II study of belinostat. Methods: We evaluated the tumor responses of 25 patients with thymic cancer using computed tomography-based RECIST, World Health Organization (WHO), modified RECIST, and volumetrics. As a control, we assessed 37 patients with non-small cell lung cancer (NSCLC) with RECIST and volumetrics. Results: Agreement analyses in 23 patients with thymic cancer at the time of RECIST-determined progressive disease (PD) compared volumetrics with RECIST, modified RECIST, and WHO criteria. Use of volumetrics was associated with 22% discordance compared with RECIST, 15% versus modified RECIST, and 22% versus WHO criteria. Volumetrics revealed PD 72 days earlier than RECIST (p = 0.016). In another cohort of 35 patients with NSCLC, there was 9% discordance between volumetrics and RECIST at the time of PD. Volumetrics demonstrated PD 32 days earlier than RECIST in NSCLC (p = 0.0078). Conclusions: Our study suggests that volumetrics might improve detection of PD. Prospective evaluation of this technique in a larger series of patients with thymic malignancies will be required.

First-line treatment of metastatic melanoma: role of nivolumab

Historically, the median overall survival of metastatic melanoma patients was less than 1 year and long-term survivors were rare. Recent advances in therapies have dramatically shifted this landscape with increased survival rates and the real possibility that long-term disease control is achievable. Advances in immune modulators, including cytotoxic T-lymphocyte antigen-4 and programmed death-1 based treatments, have been an integral part of this success. In this article, we review previous and recent therapeutic developments for metastatic melanoma patients. We discuss advances in immunotherapy while focusing on the use of nivolumab alone and in combination with other agents, including ipilimumab in advanced melanoma. One major goal in melanoma research is to optimize combination strategies allowing for more patients to experience benefit while minimizing toxicity. A better understanding of the optimal sequencing, combinations, and mechanisms underlying the development of resistance may provide evidence for rational clinical trial designs of novel immunotherapy strategies in melanoma and other cancer subtypes.

Early Stage HER2-Positive Breast Cancers Not Achieving a pCR From Neoadjuvant Trastuzumab- or Pertuzumab-Based Regimens Have an Immunosuppressive Phenotype

Background Stromal tumor‐infiltrating lymphocytes (TILs) might predict pathologic complete response (pCR) in patients with HER2‐positive (HER2+) breast cancer treated with trastuzumab (H). Docetaxel (T), carboplatin (C), H, and pertuzumab (P) have immune‐modulating effects. Pre‐ and post‐treatment immune biomarkers in cancers treated with neoadjuvant TCH with or without P are lacking. In this study we quantified baseline and changes in TILs, cluster of differentiation (CD) 4+, CD8+, FoxP3+, and PD‐L1+ cells using immunohistochemistry (IHC) and quantified productive T‐cell receptor &bgr; (TCR&bgr;) rearrangements and TCR&bgr; clonality using next‐generation sequencing (NGS) in 30 HER2+ breast cancer tissues treated with neoadjuvant H with or without P regimens. Materials and Methods Thirty pre‐ and post‐neoadjuvant TCH (n = 4) or TCHP (n = 26) breast cancer tissues were identified. TILs were quantified manually using hematoxylin and eosin. CD4, CD8, FoxP3, and PD‐L1 were stained using IHC. TCR&bgr; was evaluated using NGS. Immune infiltrates were compared between pCR and non‐pCR groups using the Wilcoxon rank sum test. Results A pCR occurred in 15 (n = 15; 50%) cancers (TCH n = 2; TCHP, n = 13). Pretreatment TILs, CD4+, CD8+, FoxP3+, and PD‐L1+ cells were not associated with response (P = .42, P = .55, P = .19, P = .66, P = .87, respectively. Pretreatment productive TCR&bgr; and TCR&bgr; clonality did not predict response, P = .84 and P = .40, respectively). However, post‐treatment CD4+ and FoxP3+ cells (T‐regulatory cells) were elevated in the non‐pCR cohort (P = .042 and P = .082, respectively). Conclusion An increase in regulatory T cells in non‐pCR tissues suggests the development of an immunosuppressive phenotype. Further investigation in a larger cohort of samples is warranted to validate these findings. Micro‐Abstract Approximately 30% to 40% of HER2‐postivie (HER2+) early‐stage breast cancers do not completely respond to neoadjuvant anti‐HER2 therapy. We investigated immune cell‐associated markers in 30 early‐stage HER2+ breast cancer tissues treated with neoadjuvant TCH (docetaxel, carboplatin, and trastuzumab) with or without pertuzumab. We identified an immunosuppressive tumor immune microenvironment in cancers with residual disease. These hypothesis‐generating results should be validated in a larger cohort.

Abstract P2-04-19: Elucidating the tumor immune microenvironment phenotype in early stage untreated BRCA mutated breast cancer patients

Background: Increased stromal tumor infiltrating lymphocytes (TILs) are predictive and prognostic for improved outcomes from neoadjuvant or adjuvant chemotherapy in triple negative breast cancer. Increased tumor mutational burden may promote neoantigens causing immune system upregulation. Microsatellite instability in gastrointestinal cancer predicts for response to checkpoint inhibition and is associated with inherited cancer predisposition. The immune system response in BRCA mutated breast cancer has not been described. The purpose of this study is to assess tumor infiltrating immune cells in early stage breast cancer patients with and without BRCA gene mutations. Methods: We retrospectively investigated 124 early stage breast cancer patients with BRCA mutations (n=62, BRCA+) and without BRCA mutations (n=62, BRCA WT). The %TILs was measured manually by HE p=0.7) groups. Median %TILs in the HR+/BRCA+ (12.5, range 0-50) and HR-/BRCA+ (15, range 5-70) cohorts were not statistically different when compared to HR+/BRCA WT (10, range 5-15; p=0.4) and HR-/BRCA WT (30, range 20-60; p=0.2) cohorts, respectively. There were 2 patients with lymphocyte predominant breast cancer (n=1, HR-/BRCA+; n=1, HR-/BRCA WT). Conclusions: This is the first study to characterize TILs and a tumor immune microenvironment phenotype in early stage breast cancer patients with BRCA mutations. These results suggest harboring a BRCA mutation is not associated with increased TILs in early stage untreated breast cancer patients. This conclusion stayed true regardless of hormone receptor status. However, a trend of decreased TILs was seen in HR-/BRCA+ patients when compared to those with HR-/BRCA WT disease. Moreover, the median and range of TILs were higher in the HR+/BRCA+ group compared to the HR+/BRCA WT group. This suggests increased TILs may exist in some HR+ patients with a BRCA mutation. Further investigation of TILs and immune profiling of early stage untreated breast cancer patients with and without BRCA mutations is warranted. Citation Format: Force J, Abbott S, Broadwater G, Kimmick G, Westbrook K, Hwang S, Kauff N, Stashko I, Weinhold K, Nair S, Hyslop T, Blackwell K, Castellar E, Marcom PK. Elucidating the tumor immune microenvironment phenotype in early stage untreated BRCA mutated breast cancer patients [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P2-04-19.