Background

The rapid evolution of palliative therapeutic choices in the last few years for patients with advanced castration-resistant prostate cancer (CRPC) has resulted in a dilemma currently troubling a few other epithelial malignancies: which systemic agent to choose and at what time? In addition, which specialty specifically directs the delivery of such care--Urology or Medical Oncology--has not been clearly established.

Approach

Recognizing the lack of consensus, we propose a framework for Urology and Medical Oncology interactions that is founded on models that have succeeded in the past.

Conclusion

This approach aims to focus the care on the patient with CRPC rather than on his physicians and promises to improve patient outcomes in this disease state.

Bladder cancer is among the top ten most common cancers, with about ~380,000 new cases and ~150,000 deaths per year worldwide. Tumor relapse following chemotherapy treatment has long been a significant challenge towards completely curing cancer. We have utilized a patient-derived bladder cancer xenograft (PDX) platform to characterize molecular mechanisms that contribute to relapse following drug treatment in advanced bladder cancer. Transcriptomic profiling of bladder cancer xenograft tumors by RNA-sequencing analysis, before and after relapse, following a 21-day cisplatin/gemcitabine drug treatment regimen identified methionine adenosyltransferase 1a (MAT1A) as one of the significantly upregulated genes following drug treatment. Survey of patient tumor sections confirmed elevated levels of MAT1A in individuals who received chemotherapy. Overexpression of MAT1A in 5637 bladder cancer cells increased tolerance to gemcitabine and stalled cell proliferation rates, suggesting MAT1A upregulation as a potential mechanism by which bladder cancer cells persist in a quiescent state to evade chemotherapy.

Gemcitabine metabolites cause the termination of DNA replication and induction of apoptosis. We determined whether subtherapeutic "microdoses" of gemcitabine are incorporated into DNA at levels that correlate to drug cytotoxicity. A pair of nearly isogenic bladder cancer cell lines differing in resistance to several chemotherapy drugs were treated with various concentrations of ¹⁴C-labeled gemcitabine for 4-24 h. Drug incorporation into DNA was determined by accelerator mass spectrometry. A mechanistic analysis determined that RRM2, a DNA synthesis protein and a known resistance factor, substantially mediated gemcitabine toxicity. These results support gemcitabine levels in DNA as a potential biomarker of drug cytotoxicity.

Precision cancer medicine seeks to target the underlying genetic alterations of cancer; however, it has been challenging to use genetic profiles of individual patients in identifying the most appropriate anti-cancer drugs. This spurred the development of patient avatars; for example, patient-derived xenografts (PDXs) established in mice and used for drug exposure studies. However, PDXs are associated with high cost, long development time and low efficiency of engraftment. Herein we explored the use of microfluidic devices or microchambers as simple and low-cost means of maintaining bladder cancer cells over extended periods of times in order to study patterns of drug responsiveness and resistance. When placed into 75 µm tall microfluidic chambers, cancer cells grew as ellipsoids reaching millimeter-scale dimeters over the course of 30 days in culture. We cultured three PDX and three clinical patient specimens with 100% success rate. The turn-around time for a typical efficacy study using microchambers was less than 10 days. Importantly, PDX-derived ellipsoids in microchambers retained patterns of drug responsiveness and resistance observed in PDX mice and also exhibited in vivo-like heterogeneity of tumor responses. Overall, this study establishes microfluidic cultures of difficult-to-maintain primary cancer cells as a useful tool for precision cancer medicine.

Background

Perturbation of the CDK4/6 pathway is frequently observed in advanced bladder cancer. We investigated the potential of targeting this pathway alone or in combination with chemotherapy or immunotherapy as a therapeutic approach for the treatment of bladder cancer METHODS: The genetic alterations of the CDK4/6 pathway in bladder cancer were first analyzed with The Cancer Genome Atlas database and validated in our bladder cancer patient-derived tumor xenografts (PDXs). Bladder cancer cell lines and mice carrying PDXs with the CDK4/6 pathway perturbations were treated with a CDK4/6 inhibitor palbociclib to determine its anticancer activity and the underlying mechanisms. The combination index method was performed to assess palbociclib and gemcitabine drug-drug interactions. Syngeneic mouse bladder cancer model BBN963 was used to assess whether palbociclib could potentiate anti-PD1 immunotherapy.

Results

Of the 413 bladder cancer specimens, 79.2% harbored pertubations along the CDK4/6 pathway. Palbociclib induced G0/G1 cell cycle arrest but with minimal apoptosis in vitro. In mice carrying PDXs, palbociclib treatment reduced tumor growth and prolonged survival from 14 to 32 days compared to vehicle only controls (p = 0.0001). Palbociclib treatment was associated with a decrease in Rb phosphorylation in both cell lines and PDXs. Palbociclib and gemcitabine exhibited antagonistic cytotoxicity in vitro (CI > 3) and in vivo, but palbociclib significantly enhanced the treatment efficacy of anti-PD1 immunotherapy and induced CD8+ T lymphocyte infiltration in syngeneic mouse models.

Conclusions

The CDK4/6 pathway is feasible as a potential target for the treatment of bladder cancer, especially in combination with immunotherapy. A CDK4/6 inhibitor should not be combined with gemcitabine.

Aim

This work aimed to determine if the treatment outcomes of bladder cancer could be improved by targeting micelles that are decorated with bladder cancer-specific ligands on the surface and loaded with the chemotherapeutic drug paclitaxel.

Materials & methods

Targeting efficacy and specificity was determined with cell lines. An in vivo targeting and anti-tumor efficacy study was conducted in mice carrying patient-derived xenografts.

Results & discussion

Targeting micelles were more efficient than nontargeting micelles in delivering the drug load into bladder cancer cells both in vitro and in vivo (p < 0.05). The micelle formulation of paclitaxel was less toxic than free paclitaxel in Cremophor(®) (Sigma, MO, USA) and allowed administration of three-times the maximum tolerated dose without increasing the toxicity. Targeting micelles were more effective than the nontargeting micelles in controlling cancer growth (p = 0.0002) and prolonging overall survival (p = 0.002).

Conclusion

Targeting micelles loaded with paclitaxel offer strong potential for clinical applications in treating bladder cancer.

Our group and others have previously shown that genistein combined polysaccharide (GCP), an aglycone isoflavone-rich extract with high bioavailability and low toxicity, can inhibit prostate cancer (CaP) cell growth and survival as well as androgen receptor (AR) activity. We now elucidate the mechanism by which this may occur using LNCaP and PC-346C CaP cell lines; GCP can inhibit intracrine androgen synthesis in CaP cells. UPLC-MS/MS and qPCR analyses demonstrated that GCP can mediate a ~3-fold decrease in testosterone levels (p < 0.001) and cause decreased expression of intracrine androgen synthesis pathway enzymes (~2.5-fold decrease of 3βHSD (p < 0.001), 17βHSD (p < 0.001), CYP17A (p < 0.01), SRB1 (p < 0.0001), and StAR (p < 0.01)), respectively. Reverse-phase HPLC fractionation and bioassay identified three active GCP fractions. Subsequent NMR and LC-MS analysis of the fraction with the highest level of activity, fraction 40, identified genistein as the primary active component of GCP responsible for its anti-proliferative, pro-apoptotic, and anti-AR activity. GCP, fraction 40, and genistein all mediated at least a ~2-fold change in these biological activities relative to vehicle control (p < 0.001). Genistein caused similar decreases in the expression of 17βHSD and CYP17A (2.5-fold (p < 0.001) and 1.5-fold decrease (p < 0.01), respectively) compared to GCP, however it did not cause altered expression of the other intracrine androgen synthesis pathway enzymes; 3βHSD, SRB1, and StAR. Our combined data indicate that GCP and/or genistein may have clinical utility and that further pre-clinical studies are warranted.

Abstract Objective To determine if a human bladder cancer-specific peptide named PLZ4 can target canine bladder cancer cells. Experimental Design The binding of PLZ4 to five established canine invasive transitional cell carcinoma (TCC) cell lines and to normal canine bladder urothelial cells was determined using the whole cell binding assay and an affinitofluorescence assay. The WST-8 assay was performed to determine whether PLZ4 affected cell viability. In vivo tumor-specific homing/targeting property and biodistribution of PLZ4 was performed in a mouse xenograft model via tail vein injection and was confirmed with ex vivo imaging. Results PLZ4 exhibited high affinity and specific dose-dependent binding to canine bladder TCC cell lines, but not to normal canine urothelial cells. No significant changes in cell viability or proliferation were observed upon incubation with PLZ4. The in vivo and ex vivo optical imaging study showed that, when linked with the near-infrared fluorescent dye Cy5.5, PLZ4 substantially accumulated at the canine bladder cancer foci in the mouse xenograft model as compared to the control. Conclusions and Clinical Relevance PLZ4 can specifically bind to canine bladder cancer cells. This suggests that the preclinical studies of PLZ4 as a potential diagnostic and therapeutic agent can be performed in dogs with naturally occurring bladder cancer, and that PLZ4 can possibly be developed in the management of canine bladder cancer.

Prostate cancer is a leading cause of cancer death among men. Currently available screening test measures prostate-specific antigen (PSA) to detect prostate cancer. However, this test produces false positive values that often lead to negative biopsies. Therefore, a more reliable diagnostic tool is needed. Glycans in serum are of particular interest as around half of all proteins are glycosylated. In this study, N-linked glycans were enzymatically released by PNGase F from prostate epithelial cell lines (pRNS) expressing wild type or mutant androgen receptors and a small set of human serum samples. Released glycans were purified and partitioned into neutral and acidic components by solid phase extraction (SPE) using graphitized carbon cartridges. The SPE fractions were analyzed by matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry (MALDI FT-ICR MS). Significant changes in some high-mannose and fucosylated biantennary complex N-linked glycans were observed in the serum of prostate cancer patients.

Background

We previously developed a bladder cancer-specific ligand (PLZ4) that can specifically bind to both human and dog bladder cancer cells in vitro and in vivo. We have also developed a micelle nanocarrier drug-delivery system. Here, we assessed whether the targeting micelles decorated with PLZ4 on the surface could specifically target dog bladder cancer cells.

Materials and methods

Micelle-building monomers (ie, telodendrimers) were synthesized through conjugation of polyethylene glycol with a cholic acid cluster at one end and PLZ4 at the other, which then self-assembled in an aqueous solution to form micelles. Dog bladder cancer cell lines were used for in vitro and in vivo drug delivery studies.

Results

Compared to nontargeting micelles, targeting PLZ4 micelles (23.2 ± 8.1 nm in diameter) loaded with the imaging agent DiD and the chemotherapeutic drug paclitaxel or daunorubicin were more efficient in targeted drug delivery and more effective in cell killing in vitro. PLZ4 facilitated the uptake of micelles together with the cargo load into the target cells. We also developed an orthotopic invasive dog bladder cancer xenograft model in mice. In vivo studies with this model showed the targeting micelles were more efficient in targeted drug delivery than the free dye (14.3×; P < 0.01) and nontargeting micelles (1.5×; P < 0.05).

Conclusion

Targeting micelles decorated with PLZ4 can selectively target dog bladder cancer cells and potentially be developed as imaging and therapeutic agents in a clinical setting. Preclinical studies of targeting micelles can be performed in dogs with spontaneous bladder cancer before proceeding with studies using human patients.