PSMA redirects cell survival signaling from the MAPK to the PI3K-AKT pathways to promote the progression of prostate cancer.
Caromile LA, Dortche K, Rahman MM, Grant CL, Stoddard C, Ferrer FA, Shapiro LH.
Sci Signal. 2017 Mar 14;10(470). pii: eaag3326. doi: 10.1126/scisignal.aag3326.
The prostates that have PSMA (left) clearly have more tumor cells (purple) that have expanded the ducts to occlude the normally clear areas, perturbed the cellular organization within the ducts and disrupted the overall structure of the prostate; consistent with rapidly progressing tumors. Cells in the tumors without PSMA (right) appear to be growing more slowly since the ductal and interstitial space and structure is maintained somewhat, consistent with a less aggressive tumor.
Research in the Shapiro Laboratory is focused on understanding the physiological and pathological regulation and function of two M1 family cell surface peptidases CD13/aminopeptidase N and PSMA/glutamate carboxy-peptidase II. This interest is the result of the striking upregulation of numerous cell surface peptidases on endothelial cells in response to both angiogenic land inflammatory signals, leading to the hypothesis that these may functionally cooperate in enzymatic cascades to regulate angiogenesis, inflammation and endothelial cell function. While the angiogenic significance of proteases that cleave large proteins (such as the matrix metalloproteases) is well documented, increasing evidence supports a role for peptidases (metAP2, CD13, APA, PSMA) as angiogenic regulators as well. The fact that these enzymes metabolize small peptide substrates suggests that small molecule regulators of angiogenesis exist which have yet to be identified and whose mechanisms are unknown. Indeed, we have shown that CD13 and PSMA are potent regulators of angiogenesis individually and our investigation of their regulatory mechanisms and their possible interaction is a current focus of the laboratory. However, our investigations have identified a variety of additional functions for these multifunctional molecules, leading our studies into the areas of molecular biology, immunology, cell biology, nephrology, transplant biology, stem cell biology, biomarkers and oncology in addition to angiogenesis using mouse models of myocardial infarction, peripheral artery disease, stroke, peritonitis, diabetic nephropathy, ureteropelvic junction obstruction and prostate cancer. Our strong relationship with clinician-scientists from the Connecticut Children’s Medical Center Departments of Surgery, Urology and Nephrology has fostered a high degree of translational research by providing clinical insights and remarkable access to patient samples, allowing us to identify, investigate and potentially treat truly clinically relevant questions.
CD13- We have shown that inhibition of CD13 blocks endothelial morphogenesis and invasion and we continue to examine the molecular mechanisms responsible for its angiogenic regulatory capabilities. Recent studies from our lab suggest that CD13 regulates signal transduction pathways leading to invasion by participating in plasma membrane organization via its interaction with membrane cholesterol and the actin cytoskeleton. Further investigations have indicated that CD13 also functions as an adhesion molecule where it mediates inflammatory cell interactions as well as endothelial/extracellular matrix interactions in a signal-transduction-dependent manner, which has strong implications for regulation of inflammatory leukocyte trafficking, cell-matrix adhesion and angiogenic cell invasion. Moreover, CD13 is expressed on embryonic and adult pluripotent stem cells and may play a role in stem cell trafficking, self-renewal and function as well. Finally, high levels of CD13 are found in the brush border lining the renal proximal tubules where we have found it regulates urinary albumin uptake in response to kidney damage resulting from diabetes. Furthermore, hypothesizing that since the proximal tubules are damaged early in renal disease, brush border proteins will be shed into the urine of pediatric patients with ureteral obstruction. To this end, we have identified a potential panel of brush border M1 enzymes, including PSMA and CD13, that may be potential biomarkers of early renal damage. Finally, we have produced a conditional CD13 knockout mouse and are actively characterizing CD13’s contribution to various physiologic and pathologic processes by specifically inactivating the gene in specific tissues.
PSMA- This M1 cell surface peptidase is also upregulated on angiogenic vasculature and its expression is dramatically increased as prostate cancer (PCa) advances, suggesting it contributes to angiogenesis and PCa progression. We have shown that this peptidase also regulates cell signaling, albeit by an apparently different mechanism than CD13. Investigation of PSMA’s regulation of endothelial cell adhesion led to the very interesting discovery that PSMA is a component of a complex regulatory loop that controls integrin signaling and PAK1 activation, thus affecting angiogenesis. Invasion studies with PSMA-null cells showed that PSMA regulates cell invasion by controlling integrin activation. These studies predicted that an extracellular-matrix-derived, small molecule PSMA substrate exists that superactivates beta1 integrins, thus regulating angiogenesis and cell invasion. Indeed, we have demonstrated that digestion of the matrix by PSMA generates small peptides that enhance endothelial cell adhesion and migration in vitro and angiogenesis in vivo via integrin α6β1 and focal adhesion kinase. Together, our results reveal a novel mechanism of PSMA activation of angiogenesis by participating in a proteolytic cascade that processes the matrix to facilitate angiogenesis.
The marked increase in PSMA expression in prostate carcinomas correlates negatively with patient prognosis, suggesting that this abundantly expressed prognostic marker contributes to PCa progression and/or metastasis. However, functional confirmation of such a role remains elusive. Recently, we have demonstrated that PSMA expression on the prostate tumor epithelium promotes tumor progression in vivo, where tumors lacking PSMA are markedly less aggressive. Understanding the functional contribution of PSMA to PCa progression and further investigation of the mechanisms of PSMA regulation of prostate tumor metastasis may lead to future novel, more effective and specifically targeted anti-cancer therapies.
- Immune Evasion in Embryonic Stem Cell-based Tissue Repair and Transplantation- Shapiro and Finck Labs
In this project we will identify CD13 as a potential target for modulating transplant immunity and elucidate the mechanisms of CD13- dependent immune evasion by investigating interactions between donor and host cells. We propose to investigate the following:
- Determine the mechanisms by which loss of CD13 circumvents immune detection of graft tissue
- Generate viable pulmonary cell types using CD13KO mESCs
- Determine if engraftment of these cells is enhanced in the lungs of immune-competent mice
- Novel biomarkers of early renal injury in children with congenital anomalies of the urinary tract- Shapiro and Ferrer labs.
Reliable biomarkers of tissue damage are invaluable tools for assessing injury and treatment efficacy. We propose to establish a panel of biomarkers specifically for patients with confirmed congenital ureteral obstruction based on the progressive course of renal damage and provide a valuable clinical tool. We will investigate the following:
- Identify a reliable panel of biomarker proteins by determining the relationship between urinary brush border protein levels and early stage renal damage in a murine model of obstruction
- Determine the utility of our biomarker panel in human urine samples obtained from pediatric patients with confirmed or suspected ureteral obstruction
- CD13 regulation of albuminuria in response to renal damage-
Shapiro lab and CCMC Dept of Nephrology
Chronic kidney disease is the slow deterioration of renal function resulting in end-stage renal failure. We propose to uncover the mechanisms underlying excess protein in the urine following kidney damage due to aberrant protein uptake and address the utility of a panel of proximal tubule proteins as biomarkers of early renal damage in pediatric patients. We will explore the following:
- Determine the molecular mechanisms by which CD13 contributes to progressive tubular disease in response to glomerular damage
- Evaluate the potential of CD13 as a therapeutic target in chronic kidney disease
- Establish a panel of brush border proteins as biomarkers for diabetic nephropathy
- CD13 in Inflammation- Shapiro lab
This project focuses on the CD13 endocytic mechanisms specifically dictating responses of myeloid cells of the innate immune system. We will study the contributions of CD13 in response to ischemic tissue damage and the mechanism by which CD13 regulates TLR4 signal transduction in response to molecules released by sterile tissue injury. We will investigate the following:
- Mechanisms by which CD13 regulates receptor-mediated endocytosis in myeloid and dendritic cells
- Contribution of CD13-dependent immune responses to impaired healing following femoral artery dissection
- Structure/function analysis of CD13’s signaling and adhesion functions using chimeric mouse/human molecules.
- Molecular mechanisms promoting prostate cancer progression by PSMA
- To determine the molecular mechanisms by which PSMA regulates signal transduction and expression of receptor tyrosine kinases and the androgen receptor to promote tumor progression and hormone-independence in prostate tumor cells.
- Evaluate human prostate tumor samples to investigate the relationship between PTEN inactivation/mutation and PSMA expression levels.
- PSMA regulation of prostate cancer metastasis.
- To determine the mechanisms responsible for decreased metastasis in tumors lacking PSMA expression and PSMA’s modulation of integrin activation to facilitate prostate tumor metastasis.
Current Collaborations and Group Projects
The Shapiro lab is part of an extremely collaborative group of scientists and clinician-scientists focusing on the biology and therapeutic targeting of pediatric diseases as outlined above. In addition, Dr. Shapiro is organizing a group of scientists and clinician scientists to submit a Program Project Group grant application, “Tissue and Organ-Specific Inflammatory Responses to Injury”, directed toward dissecting the molecular mechanisms of ischemic injury and inflammation in various tissue beds including heart, brain and peripheral muscle. Investigators include faculty from the Departments of Neuroscience, Cardiology, Vascular Biology, Immunology and Quantitative Medicine.