title: Investigation of the Lack of Angiogenesis in the Formation of Lymph Node Metastases.
abstract: BACKGROUND
  To date, antiangiogenic therapy has failed to improve overall survival in cancer patients when used in the adjuvant setting (local-regional disease with no detectable systemic metastasis). The presence of lymph node metastases worsens prognosis, however their reliance on angiogenesis for growth has not been reported.
  
  
  METHODS
  Here, we introduce a novel chronic lymph node window (CLNW) model to facilitate new discoveries in the growth and spread of lymph node metastases. We use the CLNW in multiple models of spontaneous lymphatic metastases in mice to study the vasculature of metastatic lymph nodes (n = 9-12). We further test our results in patient samples (n = 20 colon cancer patients; n = 20 head and neck cancer patients). Finally, we test the ability of antiangiogenic therapy to inhibit metastatic growth in the CLNW. All statistical tests were two-sided.
  
  
  RESULTS
  Using the CLNW, we reveal the surprising lack of sprouting angiogenesis during metastatic growth, despite the presence of hypoxia in some lesions. Treatment with two different antiangiogenic therapies showed no effect on the growth or vascular density of lymph node metastases (day 10: untreated mean = 1.2%, 95% confidence interval [CI] = 0.7% to 1.7%; control mean = 0.7%, 95% CI = 0.1% to 1.3%; DC101 mean = 0.4%, 95% CI = 0.0% to 3.3%; sunitinib mean = 0.5%, 95% CI = 0.0% to 1.0%, analysis of variance P = .34). We confirmed these findings in clinical specimens, including the lack of reduction in blood vessel density in lymph node metastases in patients treated with bevacizumab (no bevacizumab group mean = 257 vessels/mm(2), 95% CI = 149 to 365 vessels/mm(2); bevacizumab group mean = 327 vessels/mm(2), 95% CI = 140 to 514 vessels/mm(2), P = .78).
  
  
  CONCLUSIONS
  We provide preclinical and clinical evidence that sprouting angiogenesis does not occur during the growth of lymph node metastases, and thus reveals a new mechanism of treatment resistance to antiangiogenic therapy in adjuvant settings. The targets of clinically approved angiogenesis inhibitors are not active during early cancer progression in the lymph node, suggesting that inhibitors of sprouting angiogenesis as a class will not be effective in treating lymph node metastases.
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Han-Sin
authors.initials: HS
authors.lastname: Jeong
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Dennis
authors.initials: D
authors.lastname: Jones
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Shan
authors.initials: S
authors.lastname: Liao
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Wattson
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Cheryl H
authors.initials: CH
authors.lastname: Cui
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Dan G
authors.initials: DG
authors.lastname: Duda
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Willett
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL).
authors.forename: Rakesh K
authors.initials: RK
authors.lastname: Jain
authors.affiliation: Edwin L. Steele Laboratories, Department of Radiation Oncology, MGH Cancer Center, Massachusetts General Hospital and Harvard Medical School, Boston, MA (HSJ, DJ, SL, DAW, CC, DGD, RKJ, TPP); Department of Otorhinolaryngology-Head and Neck Surgery, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, South Korea (HSJ); Department of Radiation Oncology, Duke University Medical Center, Durham, NC (CGW).Current affiliation: Department of Microbiology, Immunology and Infectious Diseases, University of Calgary, Calgary, Alberta, Canada (SL). tpadera@steele.mgh.harvard.edu.
authors.forename: Timothy P
authors.initials: TP
authors.lastname: Padera
mesh_terms: D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology; D000068258:Bevacizumab; D003110:Colonic Neoplasms / Q000473:pathology*; D006258:Head and Neck Neoplasms / Q000473:pathology*; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D008198:Lymph Nodes / Q000187:drug effects / Q000473:pathology*; D008207:Lymphatic Metastasis / Q000517:prevention & control; D051379:Mice; D009389:Neovascularization, Pathologic / Q000188:drug therapy*; D011758:Pyrroles / Q000494:pharmacology; D000077210:Sunitinib
chemical_list: D020533:Angiogenesis Inhibitors; D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; C511761:DC101 monoclonal antibody; D007211:Indoles; D011758:Pyrroles; D000068258:Bevacizumab; D000077210:Sunitinib