Mechanisms of Bladder Cancer Progression

描述(由申请人提供)：识别调节膀胱癌(BCA)进展的“分子决定因素”可以改善BCA患者的治疗和复发监测。透明质酸(HA)是一种糖胺多糖，可促进肿瘤转移。透明质酸酶(Haase)是一种能将透明质酸降解成血管生成片段的酶。HYAL1是膀胱肿瘤(BT)细胞表达的主要HAase。它调节BCA在体外和Bt异种移植瘤中的生长和侵袭。虽然HYAL1野生型(Wt)具有酶活性并且仅在高级别的BCA中表达，但5个HYAL1变异体在正常和低级别的BCA组织中不具有酶活性并且表达。在Bt组织中，肿瘤细胞和间质都产生HA，而HAS1型HA合成酶仅在Bt细胞中表达。尿HA和HAase测定(HA-HASE试验)对BCA的检测具有较高的准确性。这项建议旨在研究HYAL1、HYAL1剪接变异体和HAS1在BCA进展中的治疗和预后潜力。此外，在一项多中心试验中，将评估HA-HAASE试验是否可以单独或与其他尿液试验一起用于监测BCA的复发。 为了确定HYAL1在BCA生长和进展中的作用(S)，将在Bt异种移植中测试抗HAase治疗的有效性，在使用病毒系统输送HYAL1反义cDNA后或通过HAase抑制剂治疗。将通过分析细胞周期调节因子、基质降解酶和血管生成因子的变化来研究HYAL1的作用机制(目标1)。通过将HYAL1wt表达的Bt细胞与HYAL1剪接变异体cDNAs共转染Bt细胞，检测HYAL1变异体对BCA生长和侵袭的可能中和作用。Bt组织中HYAL1变异体的差异表达将与Bt预后相关(目标2)。HAS1在Bt生长和进展中的作用将通过使用HAS1正义和HAS1反义构建体转染表达HYAL1或被阻断产生HYAL1的BCA细胞来评估。Bt组织中HAS1及其变异体(HAS1v)的表达将与BCA的预后相关(Aim 3)。在一项多中心试验中，将对HA-HASE检测、尿细胞学、BTA-Stat、NMP22检测的实用性进行单独和联合检测，以确保监测100至150名BCA患者肿瘤复发的精确度。结果将与临床结果进行比较(目标4)。 这项拟议的研究将揭示HA、HYAL1和相关分子在BT进展中的功能、治疗和/或预后潜力。此外，它将确定HA-HAASE检测或其与其他检测相结合是否可以准确地监测BCA的复发。

DESCRIPTION (provided by applicant): Identification of "molecular determinants" that regulate bladder cancer (BCa) progression could improve treatment and recurrence monitoring for BCa patients. Hyaluronic acid (HA) is a glycosaminoglycan that promotes tumor metastasis. Hyaluronidase (HAase) is an enzyme that degrades HA into angiogenic fragments. HYAL1 is the major HAase expressed in bladder tumor (BT) cells. It regulates BCa growth and invasion both in vitro and in BT xenografts. While HYAL1 wild type (wt) is enzymatically active and is exclusively expressed in high-grade BCa, 5 HYAL1 variants are enzymatically inactive and expressed in normal and low-grade BCa tissues. In BT tissues, both tumor cells and the stroma produce HA, however, HAS1 type HA-synthase is exclusively expressed in BT cells. The measurement of urinary HA and HAase levels (HA-HAase test) has high accuracy in detecting BCa. This proposal is designed to investigate the therapeutic and prognostic potentials of HYAL1, HYAL1 splice variants, and HAS1 in BCa progression. Furthermore, in a multi-center trial whether the HA-HAase test, either alone or in combination with other urine tests, can be used for monitoring BCa recurrence will be evaluated. To define HYAL1 function(s) in BCa growth and progression, the efficacy of anti-HAase therapy will be tested in BT xenografts, following delivery of HYAL1-antisense cDNA using a viral system or by treatment with a HAase inhibitor. The mechanism of HYAL1 action will be examined by analyzing alterations in cell cycle regulators, matrix degrading enzymes, and angiogenic factors (Aim 1). A possible neutralizing effect of HYAL1 variants on BCa growth and invasion will be examined by cDNA transfection of HYAL1wt expressing BT cells with HYAL1 splice variant cDNAs. Differential expression of HYAL1 variants in BT tissues will be correlated with BT prognosis (Aim 2). HAS1 function in BT growth and progression will be evaluated by transfecting BCa cells which either express or are blocked in HYAL1 production, using HAS1-sense and HAS1-antisense constructs. Expression of HAS1 and its variant (HAS1v) in BT tissues will be correlated with BCa prognosis (Aim 3). In a multi-center trial, the utility of the HA-HAase test, urine cytology, BTA-Stat, NMP22 tests, individually and in combination, will be examined for precision to monitor tumor recurrence in 100 to 150 BCa patients. The results will be compared to clinical findings (Aim 4). The proposed study will reveal the function, therapeutic and/or prognostic potentials of the HA, HYAL1 and related molecules in BT progression. Furthermore, it will establish whether the HA-HAase test or its combination with other tests can precisely monitor BCa recurrence.