Ralph A. Reisfeld, PhD: In Memoriam (1926–2020)

Cancer Research · 2021

• History
• Classics
• Gerontology

The sad news that Ralph A. Reisfeld, PhD, passed away on December 6, 2020 reached us early this year, and we would like to reflect on Ralph's long career and distinctive contribution to biomedical science and in particular to the field of cancer research. Ralph was well known for his work in biochemistry and tumor immunology and was a role model to many of his fellows and friends worldwide.Ralph was born on April 23, 1926 in Stuttgart Germany. He was the only son of Alfred and Erna Reisfeld, who were Jewish. In 1938, Ralph and his parents escaped to safety in Switzerland several days before “Kristallnacht.” For Ralph, life as a refugee was difficult, but he was grateful to the Swiss for their shelter because most of Ralph's family were killed by the National Socialists in concentration camps in Germany and Poland. Underlining his forgiving personality, Ralph never forgot his German roots, and throughout his career he made a concerted effort to develop professional and social ties with scientists from Germany. In his laboratories, he sponsored, trained, and mentored many young German scientists and young scientists from all over the world. He also never lost his love for history, which was a common subject of conversations with him.In 1947, Ralph was sponsored by the Schlesinger family of New York to immigrate to America. He had no resources, but he was anxious to begin a new life. During the ship voyage, he won $50 in a lottery, an event he took as a good omen for the future. After arriving in New York, Ralph went to work on a dairy farm in Vermont and Long Island to earn money for college. The very long days and the work ethic that farming life required stayed with him throughout his highly productive life.In 1948, Ralph began college at Rutgers University (New Brunswick, NJ), and received a Bachelor of Science Degree in Agriculture in 1952. As a student at Rutgers University, he was invited to visit with Albert Einstein in his home. They had both attended the same high school in Zurich. He considered his meeting with Einstein to be a life changing event, as his discussions with Einstein prompted him to become a scientist instead of studying history.In 1957, Ralph received a PhD in biochemistry from Ohio State University (Columbus, OH). He began his scientific career working in endocrinology at the NCI, at the NIH, in Maryland. At the NCI, he first purified and characterized human chorionic gonadotropin. A few years later, Ralph moved to Merck Sharp & Dohme in New Jersey to continue pursuing his interest in the biochemical characterization of pituitary hormones. In 1962, at Merck, he adapted PAGE and cellulose ion-exchange chromatography to this task and published an article in Nature that is still one of the most cited publications in biomedical research.In 1963, Ralph returned to the NIH as a staff scientist at the Laboratory of Immunology at the National Institute of Allergy and Infectious Diseases (Bethesda, MD), where he pioneered the characterization of immunoglobulin light chains and human histocompatibility antigens. These achievements were recognized worldwide, and his publications are listed among those that were most frequently cited in biomedical research in the following decades.In 1970, Ralph accepted the position of professor and member at the Scripps Clinic and Research Foundation (La Jolla, CA), where he continued researching serologic and molecular characterization of human MHC antigens, resulting in more than 100 publications from 1970 to 1980. Ralph was among a small group of investigators who first recognized β2-microglobulin as the light chain of MHC class I antigens. This molecular characterization of MHC facilitated the understanding of the rejection of transplanted organs.In 1976, Ralph began to work in cancer research and accomplished the biochemical and functional characterization of several melanoma- and neuroblastoma-associated antigens. He recognized their potential as therapeutic targets and developed an array of mAbs for this purpose. Ralph's group was one of the first in the world to embrace mAb technology. He supported the pioneering work of his post-doctoral fellows and associates in establishing techniques that would much later become mainstream and have tremendous impact in translation to the clinic. Several of his trainees moved to early biotechnology or pharmaceutical companies eager to develop mAbs.He was the first to characterize a melanoma-associated chondroitin sulfate proteoglycan, and demonstrated the eradication of established human melanoma and neuroblastoma tumors in nude mice by antibody-directed effector cells. The discovery of disialogangliosides, GD2 and GD3, as coreceptors of certain integrins added an important biological function to target structures previously thought to be biologically inert. In 1984, the first monoclonal anti-GD2 antibody was generated in Ralph's laboratory. This pioneering work laid the foundation for an arsenal of immunotherapeutics that were subsequently developed and are now critical to cancer treatment worldwide.In the following decade, Ralph's research focused on the development of anti-GD2 antibody constructs through protein engineering, which fostered the generation of humanized versions, antibody–drug conjugates, radio-immunoconjugates, and antibody-cytokine fusion proteins (immunocytokines). In his laboratory, these anti-GD2 constructs were tested in various preclinical cancer models to determine efficacy and mechanism of immunologic response. His findings provided the foundation for the clinical development of the anti-ganglioside GD2 antibody ch14.18. In 1989, at a time when there were no guidelines in place for antibodies generated by recombinant DNA technology, ch14.18 was the first human mouse chimeric antibody submitted to the FDA under an investigational new drug application.Through the resourceful identification of various vendors necessary to provide the required data, Ralph played a key role in the compilation of the chemistry, manufacturing, and controls package that was critical to enabling phase I/II clinical trials of ch14.18. Accordingly, ch14.18 was tested in phase III clinical trials by major cooperative pediatric oncology groups in the United States and Europe and was found to have efficacy in children with high-risk neuroblastoma. Today, ch14.18 is an FDA- and European Medicines Agency–approved drug that helps children worldwide to survive this disease. For many years, Ralph proudly displayed in his office the picture of a young boy who was one of the early recipients of ch14.18 and had his cancer go into remission.Ralph's excitement for both basic discovery and clinical application was evident to all that were privileged to meet and work with him; in his later years, he often noted how grateful he was that some of his work had an impact on treating cancer and helping children with neuroblastoma. His exemplary ability to bridge the gap between basic science and clinical application motivated his colleagues for scientific cooperation and demonstrated the tremendous translational value of basic scientific research.Ralph published more than 400 articles and received two 7-year Outstanding Investigator Awards from the NCI, underscoring his outstanding and long-lasting research achievements. He belonged to many professional societies, including the American Association for Cancer Research, to which he has been a member since 1976. He was also a member of many research grant committees, advisory groups, and editorial boards including Cancer Research and Clinical Cancer Research. In 2012, at the age of 86, and after more than 50 years in scientific research, Ralph retired from The Scripps Research Institute as Professor Emeritus.As Professor Emeritus, with his enormous experience, extensive network, and outstanding mentoring skills, Ralph continued to tutor young scientists and to support basic research and clinical scientists with their careers.Ralph frequently stated that the career achievement that he was most proud of was training and mentoring well over 100 young scientists and postdoctoral fellows, many of whom went on to conduct outstanding scientific research, to achieve Nobel Prize recognition, and to lead many prominent private companies and scientific institutions. Ralph nurtured the talent of his students and peers and always encouraged scientists to build their own ideas. He was guided by the Churchill motto, “we make a life with what we give,” and what a magnificent life Ralph gave to science.Ralph is survived by his daughter Deborah, son Daniel, daughter-in-law Lynn, and grandchildren Andrew and Lauren. His beloved wife Louise, to whom he was married for 57 years, died 7 years ago. Ralph and Louise shared a passion for classical music and loved visiting Vienna. In their later years, they made annual December trips to Vienna to attend the opera and to visit the Christmas market.Ralph was not only an outstanding scientist and enthusiastic personality, he was a lifelong and supportive friend to his students, colleagues, and collaborators. We will miss him dearly and will never forget his sense of humor, charm, and passion for science.

Silver nanospheres are cytotoxic and genotoxic to fish cells

Aquatic Toxicology · 2009-12-04 · 218 citations

Development of fragment-specific osteopontin antibodies and ELISA for quantification in human metastatic breast cancer

BMC Cancer · 2008-01-31 · 33 citations

BACKGROUND: Osteopontin (OPN) is associated with human cancers, and circulating blood OPN may have diagnostic or prognostic value in clinical oncology. METHODS: To evaluate OPN as a cancer biomarker, we generated and characterized five novel mouse monoclonal antibodies against the human full-length OPN (fl-OPN). Epitopes recognized by four antibodies (2C5, 2F10, 2H9, and 2E11) map to N-terminal OPN (aa1-166); one (1F11) maps to C-terminal OPN (aa167-314). These antibodies recognize recombinant and native OPN by ELISA and immunoblot, cross reacting with human and mouse OPN. Two of these novel antibodies (2F10 and 1F11) were used to develop a quantitative enzyme linked immunosorbent assay (ELISA) for fl-OPN. RESULTS: In comparison with commercially available ELISAs, our assay had high accuracy in measuring fl-OPN standards, and high sensitivity. Specifically, our ELISA has a linear dose response between 0.078 ng/ml-10 ng/ml, with a sensitivity of 13.9 pg/ml. We utilized this assay to quantify fl-OPN in the plasma of healthy volunteers in comparison with patients with metastatic breast cancer. The average circulating plasma fl-OPN in healthy volunteers was 1.2 ng/ml, compared to 4.76 ng/ml in patients with metastatic breast cancer (p = 0.0042). Although the increase in fl-OPN in cancer patients is consistent with previous studies, the measured quantity varied greatly between all existing fl-OPN ELISAs. CONCLUSION: Because OPN is a complex molecule with diversity from alternative splicing, post-translational modification, extracellular proteolytic modification, and participation in protein complexes, we suggest that further understanding of specific isoform recognition of multiple OPN species is essential for future studies of OPN biomarker utility.

Beta-2-Microglobulin Synthesis Is Increased during Activation of Human Monocytes

Blood Purification · 2008-10-29 · 18 citations

We have investigated the regulation of β<sub>2</sub>-microglobulin (β<sub>2</sub>-M) synthesis by monocytes. Recent interest in β<sub>2</sub>-M has developed since the discovery that this protein forms amyloid fibrils in patients undergoing long-term, chronic hemodialysis. The β<sub>2</sub>-M amyloid-osis is linked to the greatly elevated levels of monomeric β<sub>2</sub>-M in their circulation. Since factors that govern β<sub>2</sub>-M release from plasma membranes are not known, we endeavored to evaluate β<sub>2</sub>-M release during monocyte activation. Utilizing a human monocyte-like cell line, U937, we studied the effect of bacterial toxin stimulation on levels of membrane, cell surface, and supernatant β<sub>2</sub>-M. We now present a novel method to purify β<sub>2</sub>-M, a solid-phase radioimmunoassay to measure soluble β<sub>2</sub>-M, and an ELISA to measure membrane β<sub>2</sub>-M. Using these methods we found that the levels of β<sub>2</sub>-M in the cell membrane or on the cell surface did not change during monocyte activation. However, activation did induce a significant increase in the concentration of β<sub>2</sub>-M in monocyte supernatants, indicating that β<sub>2</sub>-M synthesis by monocytes is increased during monocyte activation. These results suggest that monocyte activation by hemodialysis membranes may be a contributing factor to the observed increase in circulating β<sub>2</sub>-M levels.

Announcement

˜The œNephron journals/Nephron journals · 2008-12-10

Hemodialysis-Related Induction of Beta-2-Microglobulin and Interleukin-1 Synthesis and Release by Mononuclear Phagocytes

˜The œNephron journals/Nephron journals · 2008-12-10 · 54 citations

We have investigated beta 2-microglobulin (beta 2M) synthesis and release by blood leukocytes and isolated mononuclear phagocytes. Recent interest in beta 2M has developed since the discovery that this protein forms amyloid fibrils in patients undergoing long-term, chronic hemodialysis and that these patients have greatly elevated levels of monomeric beta 2M in their circulation. Since hemodialysis-related factors that increase beta 2M production are unknown, we evaluated beta 2M production by freshly prepared leukocytes and monocyte-derived macrophages under a variety of conditions. We utilized a novel enzyme-linked immunoabsorbant assay to quantitate beta 2M concentrations, and monitored interleukin-1 and beta 2M synthesis by immunoprecipitation. Incubation of leukocytes with Cuprophan or Hemophan does not increase beta 2M release. Adherence of macrophages onto polystyrene or Cuprophan membranes induces neither interleukin-1 nor beta 2M synthesis or release. In contrast, interaction of macrophages with lipopolysaccharide, gamma-interferon, tumor necrosis factor, or interleukin-1 induces synthesis and release of beta 2 M, indicating that beta 2 M synthesis is increased during macrophage activation. Exposing leukocytes or macrophages to changes in osmotic or oncotic pressure induces a rapid release of beta 2M and interleukin-1 into the cellular medium. These results suggest that during hemodialysis, beta 2M production is more likely to result from endotoxin contamination, or osmotic and oncotic changes, rather than direct interaction of mononuclear phagocytes with Cuprophan membranes.

Widespread Delta-Like-1 Expression in Normal Adult Mouse Tissue and Injured Endothelium Is Reflected by Expression of the Dll1<sup>LacZ</sup> Locus

Journal of Vascular Research · 2007-09-26 · 13 citations

BACKGROUND: Our study characterizes Delta-like 1 (Dll1) in the adult mouse, particularly in normal versus injured vasculature, with the aid of the transgenic Dll1(LacZ) line. METHODS: Normal mouse adult tissues or those from the Dll1(LacZ) reporter line were analyzed for Dll1 expression and promoter activity. Vascular tissue was analyzed before and after carotid artery ligation. RESULTS: In wild-type mice, Dll1 transcript expression was widespread. Similarly, the Dll1(LacZ) reporter had beta-galactosidase activity detectable in the cerebellum, cerebrum, spinal cord, liver, lung and cornea, although the normal adult vasculature had no reporter expression. Following arterial ligation, there was acute induction of Dll1(LacZ) reporter expression, both in the ligated left carotid artery, and the uninjured right contralateral artery. Expression returned to low/undetectable levels 4-10 days after arterial ligation. CONCLUSION: The expression of Dll1 in the adult mouse is more widespread than previously realized, although not in resting large arteries in the adult mouse. Following arterial injury, Dll1 promoter activity is induced selectively in the endothelial cells of both the injured artery and the contralateral uninjured artery. Our results show that while overall expression in the adult mouse is widespread, Dll1 may be selectively expressed in the endothelium of injured vasculature, similar to the endothelial-restricted expression of Dll4.

Membership of the ESM

Journal of Vascular Research · 2007-09-26

Announcement

Journal of Vascular Research · 2007-09-26

Prevention of TGF-β-induced apoptosis by interlukin-4 through Akt activation and p70S6K survival signaling pathways

APOPTOSIS · 2007-07-12 · 17 citations