Lynn T. Dengel

Immunotype and Immunohistologic Characteristics of Tumor-Infiltrating Immune Cells Are Associated with Clinical Outcome in Metastatic Melanoma

Immune cells infiltrating the microenvironment of melanoma metastases may either limit or promote tumor progression, but the characteristics that distinguish these effects are obscure. In this study, we systematically evaluated the composition and organization of immune cells that infiltrated melanoma metastases in human patients. Three histologic patterns of immune cell infiltration were identified, designated immunotypes A, B, and C. Immunotype A was characterized by no immune cell infiltrate. Immunotype B was characterized by infiltration of immune cells limited only to regions proximal to intratumoral blood vessels. Immunotype C was characterized by a diffuse immune cell infiltrate throughout a metastatic tumor. These immunotypes represented 29%, 63%, and 8% of metastases with estimated median survival periods of 15, 23, and 130 months, respectively. Notably, from immunotypes A to C, there were increasing proportions of B cells and decreasing proportions of macrophages. Overall, the predominant immune cells were T cells (53%), B cell lineage cells (33%), and macrophages (13%), with natural killer and mature dendritic cells only rarely present. Whereas higher densities of CD8(+) T cells correlated best with survival, a higher density of CD45(+) leukocytes, T cells, and B cells also correlated with increased survival. Together, our findings reveal striking differences in the immune infiltrate in melanoma metastases in patients, suggesting microenvironmental differences in immune homing receptors and ligands that affect immune cell recruitment. These findings are important, not only by revealing how the immune microenvironment can affect outcomes but also because they reveal characteristics that may help improve individualized therapy for patients with metastatic melanoma.

Effectiveness of imiquimod limited to dermal melanoma metastases, with simultaneous resistance of subcutaneous metastasis

Successful management of epithelial skin cancers with imiquimod 5% cream (Aldara®), an immunomodulatory agent, led to speculation that it may promote an immune response against melanoma. Studies, mostly case reports, have assessed the value of imiquimod as a topical treatment for dermal melanoma metastases that prove difficult to manage surgically. The precise value of imiquimod, however, in treatment of dermal and subcutaneous metastases remains unclear. A case at our institution elucidates histopathologically that subcutaneous metastases may progress despite excellent treatment of superficial dermis in the same location. In preparation for a clinical trial using imiquimod to treat patients with dermal melanoma metastases, we have treated several patients off protocol. We present a case report in which the observed changes are documented photographically and histologically. The patient experienced dramatic improvement in the locally treated dermis with concurrent regional treatment failure in the subcutaneous space. Our experience supports growing evidence that imiquimod for some provides an effective option for dermal disease. The unique histological documentation we provide regarding the differential effectiveness of imiquimod in treating various tissue components may help guide future investigations regarding optimal clinical application of imiquimod therapy for melanoma metastases.

Interferons induce CXCR3-cognate chemokine production by human metastatic melanoma.

Immune-mediated cancer regression requires tumor infiltration by antigen-specific effector T cells, but lymphocytes are commonly sparse in melanoma metastases. Activated T cells express CXCR3, whose cognate chemokines are CXCL9/MIG, CXCL10/IP-10, and CXCL11/I-TAC. Little is known about expression of these chemokines in lymph node (LN) metastases of melanoma. We evaluated whether metastatic melanoma induces these CXCR3-cognate chemokines in human LN-derived tissues. In addition, as these chemokines can be induced by interferon (IFN), we evaluated whether type I or II IFNs (IFN-α or IFN-γ, respectively) can modulate chemokine expression in an in vitro model of the human tumor microenvironment. Production of CXCL9-11 by melanoma-infiltrated nodes (MIN) was no different than tumor-free nodes; both produced less chemokine than activated LN (sentinel immunized nodes, SIN). These data suggest that melanoma infiltration into LN neither induces nor reduces CXCL9-11. Stimulation with IFN-α or IFN-γ increased production of CXCL10-11 from MIN, but not tumor-free node or SIN. IFN-γ also increased production of CXCL9 in MIN. In IFN-treated SIN, CD14+ cells were the primary source of CXCL9-11, whereas melanoma cells were the source of chemokine in MIN. Melanoma cells in MIN express IFN receptors. Consistent with these observations, multiple human melanoma lines expressed IFN receptors and produced CXCL9-11 in response to IFN treatment. Thus, melanoma infiltration of LN is insufficient to induce the production of CXCL9-11, but melanoma may be a significant source of IFN-induced chemokines. Collectively, these data suggest that IFN-α or IFN-γ may act in the tumor microenvironment to increase the chemotactic gradient for CXCR3+ T cells.

Intraoperative Imaging Guidance for Sentinel Node Biopsy in Melanoma Using a Mobile Gamma Camera

Objective:To evaluate the sensitivity and clinical utility of intraoperative mobile gamma camera (MGC) imaging in sentinel lymph node biopsy (SLNB) in melanoma. Background:The false-negative rate for SLNB for melanoma is approximately 17%, for which failure to identify the sentinel lymph node (SLN) is a major cause. Intraoperative imaging may aid in detection of SLN near the primary site, in ambiguous locations, and after excision of each SLN. The present pilot study reports outcomes with a prototype MGC designed for rapid intraoperative image acquisition. We hypothesized that intraoperative use of the MGC would be feasible and that sensitivity would be at least 90%. Methods:From April to September 2008, 20 patients underwent Tc99 sulfur colloid lymphoscintigraphy, and SLNB was performed with use of a conventional fixed gamma camera (FGC), and gamma probe followed by intraoperative MGC imaging. Sensitivity was calculated for each detection method. Intraoperative logistical challenges were scored. Cases in which MGC provided clinical benefit were recorded. Results:Sensitivity for detecting SLN basins was 97% for the FGC and 90% for the MGC. A total of 46 SLN were identified: 32 (70%) were identified as distinct hot spots by preoperative FGC imaging, 31 (67%) by preoperative MGC imaging, and 43 (93%) by MGC imaging pre- or intraoperatively. The gamma probe identified 44 (96%) independent of MGC imaging. The MGC provided defined clinical benefit as an addition to standard practice in 5 (25%) of 20 patients. Mean score for MGC logistic feasibility was 2 on a scale of 1–9 (1 = best). Conclusions:Intraoperative MGC imaging provides additional information when standard techniques fail or are ambiguous. Sensitivity is 90% and can be increased. This pilot study has identified ways to improve the usefulness of an MGC for intraoperative imaging, which holds promise for reducing false negatives of SLNB for melanoma.

Neonatal Congenital Malignant Melanoma With Lymph Node Metastasis

A Caucasian baby girl presented at birth with a dark, irregular, raised skin lesion measuring approximately 2 cm in diameter on the medial aspect of her right leg (Fig 1). There was no history of melanoma in her family and no evidence of melanoma in her mother. At two weeks of age, she underwent an incisional biopsy of the lesion at a community hospital. Histologic review showed a superficial spreading invasive malignant melanoma with Breslow thickness greater than 2.5 mm, with a positive deep margin. There was angiolymphatic invasion but no definite ulceration. She was referred to our institution, where clinical exam revealed an otherwise healthy newborn girl, a black lesion on the right leg with a central scar measuring 1.5 cm in diameter. There were small palpable inguinal nodes in each groin, possibly more prominent on the right but not clinically suspicious. One week later, a wide local excision was performed with a 1 cm margin (Fig 1) and a full thickness skin graft. Lymphoscintigraphy revealed a sentinel node in the right groin, and a sentinel lymph node biopsy was performed. Her final pathology revealed an atypical malignant melanoma arising in a nevus with congenital features with Breslow thickness of 2.58 mm and Clark’s level V (Fig 2A). Margins were widely clear. The lesion was consistent with a primary melanoma, rather than metastatic disease from maternal origin. The sentinel node was positive for metastatic melanoma, with mitotic figures (Fig 2B) and S100 staining (Fig 2C). She underwent staging studies including CT scans of her head, chest, abdomen and pelvis as well as a bone scan. These were negative for metastatic disease. One month later, completion right inguinal lymph node dissection was performed, yielding five lymph nodes without metastatic disease. Her postoperative course was complicated by a wound abscess in the right groin that was drained operatively and healed well. Her final staging, at the age of 9 weeks, was therefore Stage IIIA (T3aN1aM0). No additional treatment was performed. She has been followed to her current age of 32 months without evidence of recurrence by clinical exam, serial chest radiographs, and annual CT scans of the abdomen and pelvis. She has had an excisional biopsy of a suspicious skin lesion from her lower abdomen at age 22 months that was an atypical nevus. She has normal function of her right lower extremity without lymphedema and is healthy and developmentally appropriate (Fig 3).

Total body photography for skin cancer screening

Total body photography may aid in melanoma screening but is not widely applied due to time and cost. We hypothesized that a near‐simultaneous automated skin photo‐acquisition system would be acceptable to patients and could rapidly obtain total body images that enable visualization of pigmented skin lesions.

Intratumoral interferon-gamma increases chemokine production but fails to increase T cell infiltration of human melanoma metastases

IntroductionOptimal approaches to induce T cell infiltration of tumors are not known. Chemokines CXCL9, CXCL10, and CXCL11 support effector T cell recruitment and may be induced by IFN. This study tests the hypothesis that intratumoral administration of IFNγ will induce CXCL9–11 and will induce T cell recruitment and anti-tumor immune signatures in melanoma metastases.Patients and methodsNine eligible patients were immunized with a vaccine comprised of 12 class I MHC-restricted melanoma peptides and received IFNγ intratumorally. Effects on the tumor microenvironment were evaluated in sequential tumor biopsies. Adverse events (AEs) were recorded. T cell responses to vaccination were assessed in PBMC by IFNγ ELISPOT assay. Tumor biopsies were evaluated for immune cell infiltration, chemokine protein expression, and gene expression.ResultsVaccination and intratumoral administration of IFNγ were well tolerated. Circulating T cell responses to vaccine were detected in six of nine patients. IFNγ increased production of chemokines CXCL10, CXCL11, and CCL5 in patient tumors. Neither vaccination alone, nor the addition of IFNγ promoted immune cell infiltration or induced anti-tumor immune gene signatures.ConclusionThe melanoma vaccine induced circulating T cell responses, but it failed to infiltrate metastases, thus highlighting the need for combination strategies to support T cell infiltration. A single intratumoral injection of IFNγ induced T cell-attracting chemokines; however, it also induced secondary immune regulation that may paradoxically limit immune infiltration and effector functions. Alternate dosing strategies or additional combinatorial treatments may be needed to promote trafficking and retention of tumor-reactive T cells in melanoma metastases.

Academic or community practice? What is driving decision-making and career choices

Background: Identifying factors that impact progression of surgery trainees into academic versus non‐academic practices may permit tailoring residency experiences to promote academic careers in institutions charged with the training of future surgeon scientists. The aim of this study was to identify factors associated with progression of surgery trainees into academic versus non‐academic practice. Methods: A survey was distributed to 135 surgeons graduating from the University of Virginia residency program from 1964–2016, a single academic institution. Questions addressed practice type, research productivity, work/life balance, mentorship, and overall sentiment toward research and academic surgery. A 5‐point Likert scale measured career satisfaction and influence of factors in practice setting choice. Results: Of the 135 surveys that were electronically distributed, 69 participants responded (response rate: 51%). Of the 54 with known current practice types, 34 (63%) were academic and 20 (37%) non‐academic. Academic surgeons reported more publications by the conclusion of surgery training (56% vs 25% with >10 publications, P = .02). More academic surgeons reported >

Abstract 3827: Interferons induce CXCR3-cognate chemokine production by melanoma

100,000 in student debt at graduation (44% vs 25%, P < .05). Factors encouraging an academic career were similar for both types of surgeons, including involvement in education of trainees and access to mentorship. Both groups were discouraged from an academic practice by requirements of grant‐writing and funding responsibilities. Surgeons in academic practice were more likely to recommend surgery as a career to a current medical student (100% vs 70%, P = .001). Conclusion: This knowledge may help to tailor training experiences to promote academic careers. By supporting funding mechanisms and grant‐writing programs, while encouraging mentorship and productive research experiences, current surgical trainees may be more enthusiastic about a career in academic practice.

Short length of stay and rapid recovery to normal function after surgery for metastatic melanoma to abdominal and retroperitoneal viscera

Immune-mediated cancer regression requires tumor infiltration by Ag-specific effector T cells, but lymphocytes are commonly sparse in melanoma metastases. Activated T cells express CXCR3, whose cognate chemokines are CXCL9/MIG, CXCL10/IP-10 and CXCL11/I-TAC. Little is known about expression of these chemokines in lymph node (LN) metastases of melanoma. We evaluated whether metastatic melanoma induces these CXCR3-cognate chemokines in human LN-derived tissues. Also, because these chemokines can be induced by interferon (IFN), we evaluated whether type I or II IFNs (IFN-α or IFN-γ, respectively) can modulate chemokine expression in an in vitro model of the human tumor microenvironment. Production of CXCL9-11 by melanoma-infiltrated nodes (MIN) was no different than tumor-free nodes (TFN); both produced less than activated LN (sentinel immunized nodes, SIN). These data suggest melanoma infiltration into LN neither induces nor reduces CXCL9-11. Stimulation with IFN-α or IFN-γ increased production of CXCL10-11 from MIN, but not TFN or SIN. IFN-γ also increased production of CXCL9 in MIN. In IFN-treated MIN, the primary source of CXCL9-11 was melanoma cells themselves, whereas CD14+ cells were the source of these molecules in SIN. Melanoma cell lines expressed IFN receptors and produced CXCL9-11 in response to IFN treatment. Treatment of melanoma cell lines or primary melanomas in vitro with IFN-γ induced the migration of CXCR3+ T cells in transwell assays, whereas T cell did not significantly migrate in response to resting melanoma cells. Thus, melanoma infiltration of LN is insufficient to induce the production of CXCL9-11, but melanoma may be a significant source of IFN-induced chemokines. Collectively, these data suggest that IFN-α or IFN-γ may act in the tumor microenvironment to increase the chemotactic gradient for CXCR3+ T cells, thus enhancing the capacity of therapeutic vaccines to mediate clinical efficacy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3827.