[IF 21.7] Absin Multiplex Kit & Antibody Empower Flaxseed Lignan + PD-1 Blockade Study in Breast Cancer, Unraveling Gut Microbiota–Immune Synergy

I. Background & Rationale: Targeting immunotherapy resistance with a natural-product–microbiome axis

1. Clinical challenge: limited single-agent activity of PD-1 blockade

PD-1/PD-L1 inhibitors (PDi) yield objective responses in only a minority of breast-cancer patients; most tumours are either refractory or acquire resistance. Rational partners that can “ignite” anti-tumour immunity and reverse resistance are therefore urgently sought.

2. Working hypothesis: flaxseed lignans as an ideal partner

Flaxseed lignans (FL; enriched in secoisolariciresinol diglucoside, SDG) exhibit proven anti-cancer activity. We hypothesised that FL could potentiate PDi efficacy by re-programming the gut microbiome and/or host immunity—two determinants now recognised to govern immunotherapy responses.

3. Three-step experimental roadmap

1. ① Demonstrate intrinsic anti-tumour activity of FL in vitro and in vivo and test its dependence on the gut microbiota;
2. ② Identify the key microbial metabolite (enterolactone, ENL) and downstream host target (CD38) to establish the FL→microbiome→ENL→CD38 axis;
3. ③ Evaluate therapeutic synergy between FL and PD-1 blockade (BMS-1) while mapping concomitant changes in the microbiome (e.g., Akkermansia) and the TIME.

II. Key Findings: FL + PD-1 blockade exerts dual control via microbiome–immune crosstalk

1. FL suppresses breast-cancer progression through the gut-microbiome→ENL→CD38 axis

Core discovery: FL requires microbial conversion to enterolactone (ENL) to down-regulate CD38—a gene linked to immune evasion and PDi resistance—thereby inhibiting proliferation, migration and invasion of 4T1 and BT549 cells.

Fig.: Tumour volume/weight changes in FL-treated breast-cancer mice and impact of antibiotic depletion (original Fig. 1).

In-vivo validation: Oral FL markedly reduced tumour burden; antibiotic-mediated microbiota ablation attenuated the benefit, while ENL supplementation or faecal microbiota transplantation (FMT) from FL-treated donors restored anti-tumour efficacy.

Mechanistic proof: Genetic CD38 over-expression abolished the anti-proliferative effect of ENL and abrogated FL monotherapy activity, confirming CD38 as the critical target.

2. FL markedly potentiates PD-1 blockade

Fig.: Anti-tumour efficacy of FL combined with PD-1 inhibitor BMS-1 (original Fig. 6).

Combination benefit: FL + BMS-1 (FLcPDi) produced superior tumour growth inhibition versus either single agent, without overt toxicity (no body-weight loss).

Molecular read-out: IHC confirmed significant CD38 down-regulation in FL and FLcPDi groups, corroborating the proposed mechanism.

3. FL reshapes the gut microbiome and the tumour immune microenvironment (TIME)

Fig.: TIME composition (CyTOF) and microbiota shifts after FLcPDi (original Fig. 7).

Microbiota signature: 16S rDNA sequencing revealed a selective enrichment of Akkermansia muciniphila in FLcPDi-treated mice; gavage with live Akkermansia restored PDi sensitivity in antibiotic-pretreated animals, identifying this genus as a key synergistic taxon.

TIME reprogramming: Multiplex IHC showed increased infiltration of CD3⁺, CD4⁺ and CD8⁺ T cells and reduced immunosuppressive F4/80⁺ macrophages after FL. CyTOF further revealed expansion of memory CD4⁺ and CD8⁺ subsets and a decrease in M2-like macrophages, collectively indicating an immune-stimulatory TIME.

III. Absin Products: High-precision tools enabling mechanistic dissection

All TIME imaging and CD38 validation data were generated with Absin reagents; two flagship products underpinned the study:

1. Five-colour multiplex immunofluorescence kit (rabbit secondary) (Cat. abs50029–100T)

Application: Multiplex immunohistochemistry (mIHC) on FFPE tumour sections.

Utility: Simultaneous detection of CD3⁺ (orange), CD4⁺ (yellow), CD8⁺ (pink) and F4/80⁺ (red) cells in a single tissue slice, enabling quantitative profiling of TIME remodelling by FL.

Fig.: TIME infiltration changes after FL (original Fig. 5D-E).

Robust fluorophore stability and high specificity circumvent the throughput limitations of conventional single-colour IHC.

2. Rabbit anti-CD38 polyclonal antibody (Cat. abs136071)

Application: Chromogenic and fluorescent IHC.

Utility: Semiquantitative assessment of CD38 protein levels across treatment arms, providing histological evidence for FL-mediated CD38 down-regulation.

Fig.: CD38 IHC signal across groups (original Fig. 6D-E, I-J).

High affinity and low background deliver reliable spatial resolution of CD38 expression differences.

IV. Conclusions & Outlook: Empowering translational immuno-oncology with Absin

This study establishes a clinically translatable “natural product + immune-checkpoint blockade” paradigm and delineates a multi-level mechanism linking gut-microbiota metabolism to target modulation and TIME reprogramming. Absin’s TSA-based multiplex kits and ultra-specific antibodies served as the analytical backbone, enabling high-resolution dissection of complex immune landscapes.

As cancer immunotherapy moves toward microbiome–immune–metabolism integration, Absin will continue to provide cutting-edge tools—including expanded mIHC panels, flow-cytometry antibodies and customised conjugates—to accelerate mechanistic discoveries and their clinical translation.

References

Wu H, et al. The combination of flaxseed lignans and PD-1/PD-L1 inhibitor inhibits breast cancer growth via modulating gut microbiome and host immunity. Drug Resistance Updates. 2025;80:101222.

Products Cited in the Study

Extended Multiplex IHC Portfolio

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