AI News, Research helps explain why androgen-deprivation therapy doesn't work for many prostate cancers

Research helps explain why androgen-deprivation therapy doesn't work for many prostate cancers

'Androgen-deprivation therapy is commonly used to treat patients whose prostate cancer has spread beyond the prostate.

We have discovered a mechanism that causes progression to this aggressive form of prostate cancer, providing a new opportunity to prevent or treat lethal forms of prostate cancer,' says co-senior author David Goodrich, PhD, Professor of Oncology in the Department of Pharmacology and Therapeutics at Roswell Park.

'Importantly, these findings offer a new understanding of prostate cancer lineage plasticity, which involves the conversion of cancer cells that are dependent on a specific therapeutic target to cancer cells that are now indifferent to that target's function,' adds co-senior author Leigh Ellis, PhD, Assistant Professor of Oncology in the Department of Pharmacology and Therapeutics.

MYC regulates ductal-neuroendocrine lineage plasticity in pancreatic ductal adenocarcinoma associated with poor outcome and chemoresistance




47, KPC and KPC Myc

fl/+[28] and ROSA-LSL-Myc

30 mice have been previously described.


;Pdx1-Cre mice31 were obtained from Dr. Kerry Campbell at Fox Chase Cancer Center.

G12D;Ptf1a-CreERT;ROSA-YFP (KCY) mice26 were treated with 5 mg tamoxifen once a day for 5 consecutive days to induce acinar cell specific recombination.

2 mm3 pieces of primary pancreatic tumor tissue, resected from patients with informed written patient consent (collected in collaboration with the Brenden-Colson Center for Pancreatic Care, IRB approved, IRB00003330), were coated in Matrigel and implanted subcutaneously into the flank of 6-week-old NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice (passage 1).

One milliliter was applied via microfluidic pump to a geometrically enhanced differential immunocapture (GEDI) chip functionalized previously with capture antibodies specific to epithelial cell adhesion molecule (EpCAM).

PCA from the Beltran et al.11 and checked for expression of these genes in the TuGMP (black bars) and matched GMP circulating tumor cells (CTCs–red bars) from the single cell mouse RNA-Seq data from Ting et al.23, accession number: GSE51372.

For GSEA analysis to determine if NEPC upregulated genes were enriched in human PDA subtypes, we used the 96 PDAC ICGC RNAseq dataset from Bailey et al.24 and their designated subtypes for each sample identified, looking for enrichment of the upregulated genes in the NEPC vs.

For analysis of MYC chIP seq data in KPC tumor cells from Walz et al.28, we selected peaks with an FDR q-value of less than or equal to 0.01, annotated them with the 5 prime ends of Ref-Seq genes and then selected all genes with one or more MYC ChIP peaks within 1000 bases of their 5 prime end.

For qRT-PCR, the cycle numbers for each run were used to calculate fold change using the ddt(ct) method between each sample and the average of the control sample (set as 1) in gene expression as follows: dt(ct) = SYP cycle # - GAPDH cycle # ddt(ct) = dt(ct) – average dt(ct) of normal samples Fold Change = Power(2,ddt(ct)) Cells were homogenized in ice-cold AB-MYC lysis buffer (1 M Tris pH 7.5, 5 M NaCl, 10% Triton X-100, 10% DOC, 20% SDS, 0.5 M EDTA, 1× protease and phosphatase inhibitors (Roche, Indianapolis, Indiana)), passed through a 30 gauge needle, and cleared by centrifugation at 4 °C, 13,000xg for 10 min.

Unless otherwise stated in the figure legends, P values were calculated using a standard Student’s t test analysis (two-tailed distribution and two-sample unequal variance) to determine statistical significance as indicated in the graphs.

To determine if CK-SYP staining is an independent risk factor for poor outcome in PDA patients, we performed multivariate analysis to determine if clinical stage, grade or age significantly altered relationship between CK-SYP co-staining and days-to-recurrence.

<?xml version="1.0" encoding="UTF-8"?>TGFβ signaling limits lineage plasticity in prostate cancer

Here we show that in the background of a Pten null mutation in mouse prostate, deletion of Tgfbr2 results in increased basal cell proliferation as tumors become invasive.

Disruption of TGFß signaling either by Smad4 or Tgfbr2 deletion in the background of a prostate specific tumor initiating mutation results in rapid progression to invasive and metastatic cancer in mouse models [20, 24].

This increase appears to be due to increased basal cell numbers in these tumors following Tgfbr2 deletion, and we show that a major effect of TGFß signaling in this context is to limit proliferation of basal cells.

Analysis of gene expression data suggests that in comparison to the Pten null tumors, disruption of TGFß signaling primarily affects expression of gene sets associated with highly proliferative, aggressive cancers.

Although gene sets associated with EMT or invasion are enriched in the tumors compared to wild type prostate, it appears that such invasive gene expression signatures are already enriched by deletion of Pten alone.

Regions of micro-invasion may be detectable early, but large locally invasive cancers are not generally detected in the Pten model until 30–50 weeks of age, and metastases are extremely rare [19, 20].

Using the Krt8-CreERT2 driver here, and initiating tamoxifen treatment at 4 weeks after birth, median Pten;Tgfbr2 double mutant survival was 17–18 weeks of age, or 13–14 weeks after initiating tamoxifen treatment.

Other tamoxifen dosing regimens resulted in similar phenotypes with reduced penetrance and longer lag times, although we did not attempt to induce recombination in very old animals.

We searched extensively for basal cells in which recombination of the lineage tracing reporter could be detected early after tamoxifen treatment and did not find any evidence for recombination in Krt5 positive basal cells.

Since basal cell proliferation is so rapid in the absence of Tgfbr2, if rare basal cells underwent recombination we would expect to identify small focal regions of basal cells within the tumors, without the relatively high proportion of cells expressing both basal and luminal keratins.

If these cells arise from rare recombination events in uncommitted progenitors, or rare dual positive cells, it seems unlikely that they would become such a large proportion of the tumor given that their proliferation rate is lower than that of other cell types.

The fact that we see both dual positive and basal cells at high frequency in invasive cancers, while HGPIN is exclusively luminal suggests that both dual positive and basal cells arise from luminal cells lacking Pten and Tgfbr2.

One possibility is that relatively rare multipotent cells may be able to survive androgen ablation and allow subsequent relapse even after the majority of the tumor, which has a primarily luminal phenotype, has regressed.

Our data suggest that TGFß signaling acts to maintain the differentiated state of luminal tumor cells and that loss of this signaling pathway allows de-differentiation to an intermediate cell type, which can then further differentiate to a basal cell phenotype.

While the final differentiation to basal cells as seen in our model may not be the usual situation in human CaP, the generation of de-differentiated intermediate cells might provide insight into how these cells arise in human CaP.

Although recurrent inactivating mutations or deletions of the major components of the TGFß signaling pathways are not found in human CaP, there is evidence for reduced expression of both TGFBR1 and TGFBR2 in more advanced CaP [12–14], and reduced SMAD4 and TGFBR2 expression due to promoter methylation [9, 11].

While the mouse models used here are not ideal for addressing all questions regarding advanced human CaP, they may uncover important features of earlier stages of disease progression and metastasis.

Recent work with mouse models and xenografts has suggested that androgen ablation selects for a more de-differentiated cell type with increased lineage plasticity in CaP [40, 41, 54].

Once the recurrent disease has progressed to a more advanced stage such intermediate cells will likely have differentiated, either to a neuroendocrine phenotype [40, 41, 54], or to androgen-independent non-neuroendocrine tumors [55].

While the Krt8-CreERT2 model used here results in large numbers of de-differentiated dual positive cells, one limitation is that these tumors are overtaken by basal cells, possibly derived from the dual positive population.

We do not know if interfering with androgen signaling affects the generation of dual positive cells in this model, but this may be of interest given the possibility that anti-androgen therapy selects for de-differentiated cells in human CaP.

The frequency of post-ADT tumors with dual positive cells was more than 25% (3 of 11 samples), despite the heterogenous modes of ADT, and the variable time after treatment that the samples were isolated.

Even in a single tumor, ducts with HGPIN are purely luminal whereas adjacent regions of invasive cancer frequently contain a mix of luminal, basal and dual positive cells.

Importantly, by combining luminal specific deletion, lineage tracing and extensive analysis of basal and luminal markers, we provide evidence for a transition from luminal tumor cells to a de-differentiated invasive cell type that may be important for metastatic spread.

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