Phage Therapeutics & Enzymes (Lysins / Depolymerases) — CDMO Services

Design → GMP, without detours. Microbial-first development and manufacturing for lytic bacteriophages, phage-derived enzymes, and combination modalities—built to pass audits, survive scale, and accelerate your IND/CTA.

Why phage therapeutics and lysins/depolymerases—why now?

Antimicrobial resistance has made targeted, bio-rational approaches urgent. Lytic bacteriophages kill with exquisite specificity; phage-derived endolysins cleave peptidoglycan with single-step lethality; capsular depolymerases strip protective polysaccharides to unmask pathogens and sensitize them to immunity and antibiotics. Together, these agents promise precision where broad-spectrum drugs struggle. Regulators have clarified expectations: in the United States, clinical use of unlicensed phage products proceeds under IND with CBER/OVRR oversight; in Europe, the EMA has initiated a dedicated phage therapy quality/manufacturing concept paper to formalize phage-specific GMP and control strategies.

Mika Biologics is a microbial-specialist CDMO. We design your program end-to-end—from host-range screening and enzyme selection through GMP fermentation, downstream purification, and aseptic fill/finish—with stability and release set up to pass scrutiny. We map QTPP → CQAs → CPPs from day one and keep that line straight through PPQ.

Subtle promise: from first plaque to first patient, the path feels shorter, the risk feels lighter.

What we make

• Lytic bacteriophage drug substance (DS) and drug product (DP): single phage or cocktails, with well-defined bank strategy, identity, potency, impurity controls, and comparability. No temperate phages/lysogeny for therapeutic use; optional engineered phages handled under GMO-aware controls per EU expectations.
• Endolysins (Gram-positive focus; engineered “artilysins/innolysins” or permeabilizer-enabled variants for Gram-negatives), produced in microbial hosts, refolded if needed, polished to low endotoxin, and potency-qualified.
• Capsular depolymerases (e.g., Klebsiella, Acinetobacter): expressed and purified as recombinant enzymes or captured from phage particles, with activity verified against defined K-types/serotypes and synergy with antibiotics or serum.

We also build combination programs (e.g., phage + lysin, depolymerase + antibiotic), with study designs that generate convincing potency narratives—even post-ContraFect lessons learned.

The Mika path: host-range screening → GMP fermentation → aseptic fill/finish

Discovery & Host-Range Screening

Panel design. We curate or incorporate your clinical isolates (geographically and temporally stratified), reference strains, and resistance phenotypes (ESBL, carbapenemase, MRSA/VRSA).
Primary screens.

• Spot tests for rapid lysis signal on agar lawns (high-throughput; prone to overestimation).
• EOP (Efficiency of Plating) for quantitative infectivity: titer on each strain vs. maximal titer—our selection gold standard.
  Secondary / orthogonal readouts. 96-well growth-inhibition curves, micro-plaque assays, single-cell time-lapse (selected projects), and liquid-culture virulence metrics.

Genomic suitability. We confirm strictly lytic biology, absence of toxins/ARGs, and define termini/packaging sites; engineered phages receive GMO-aware controls aligned to the EMA concept paper’s bank and terminology proposals.

Cocktail design. We rationalize breadth with minimal redundancy: non-overlapping receptors, additive EOP coverage, escape-resistance pressure tests (serial passage). Protocols track with compassionate-use “phage matching” playbooks, adapted for regulated programs.

For enzymes. Lysin candidates undergo turbidimetric kill kinetics, zymograms, MIC/time-kill vs. planktonic & biofilm states; depolymerases get capsule-clearing assays and serum synergy tests.

Upstream Manufacturing (Phage DS) — GMP-aware

Seed & host banks. MCB/WCB for the production host (virulence-determinant-free; no prophage) and phage master/working seed with full CoAs (titer, identity, purity). Closed bioreactors with monitored DO, pH, temperature, agitation; infection MOI optimized by growth phase. CIP/SIP or validated single-use trains.

In-process controls (IPC). Growth kinetics (OD), lysis onset, PFU kinetics, residual viable host, and nuclease dosing readiness to curtail free DNA early. We document bioburden and adventitious agents.

Downstream Purification (Phage DS)

We deploy clarification → nuclease conditioning → TFF (UF/DF) → chromatography sequences that consistently separate virions from host debris, DNA, and endotoxin—without antique CsCl gradients.

• TFF (UF/DF) concentrates and buffer-exchanges with scalable shear control; we choose MWCO and flux to protect tails/capsids.
• Anion-exchange (monoliths or membranes) binds virions and separates LPS/host proteins; enzyme pretreatments can tune charge to improve resolution.
• Polishing removes carry-through aggregates and residual nucleic acids. We track recovery and log-reduce for each impurity.
• Endotoxin control strategy spans process design and acceptance criteria consistent with FDA’s Q&A and modern, non-animal BET options (USP <86> rFC/rCR) as appropriate.

Formulation preview. Early DS buffers mirror intended DP needs (ionic strength, divalent cations, protectants), smoothing the handoff to formulation.

Recombinant Enzyme Manufacturing (Lysins / Depolymerases)

Expression. E. coli for speed and isotopically typical peptides; Pichia when secretion, glyco-tuning, or protease background favors quality. Inclusion-body paths leverage redox refolds; secreted paths favor simplified DSP.
Purification. Affinity (tagged or tag-cleaved), IEX/HIC and SEC; endotoxin removal via phase-appropriate AEX/polymixin-like resins or membrane adsorbers; final polish to minimize aggregates.
Potency & stability. Robust activity assays (turbidity reduction, time-kill) and orthogonal biophysics (DSC, nanoDSF, SEC-MALS). For Gram-negative targeting, we evaluate permeabilizer co-formulation or engineered “innolysins.”

Formulation & Aseptic Fill/Finish

Phage DP. We tailor ionic/osmotic balance, pH windows, divalent cations, and excipients (e.g., sugars, amino acids) to maximize titer retention across cold chain and lyophilization; surfactants are used cautiously. Lyophilized DP can enable room-temperature field use and logistics.

Enzyme DP. Protein-centric formulation (pH 5–8; histidine/citrate/phosphate systems), aggregation control (tonicity agents), oxidation mitigation, and container-closure interactions. High-concentration syringes require viscosity mapping and glide-force studies.

Aseptic processing. We execute Grade A isolator filling (vials, PFS, cartridges) within Grade B background, with environmental/personnel monitoring to EU GMP Annex 1 (2022) expectations and current operation dates. CCIT and media fills are standard.

Analytics & release: what “good” looks like

For phage DS/DP

• Identity & purity: Whole-genome confirmation; restriction/coverage fingerprints; residual host DNA (qPCR/ddPCR), residual protein (ELISA), and adventitious agents panel.
• Potency: Plaque assay PFU/mL (validated DLA overlay), EOP on panel, liquid-culture kill kinetics.
• Safety: Endotoxin by USP <85> or USP <86 recombinant methods (BET), Sterility by USP <71 (method suitability documented).
• Other CQAs: pH, osmolality, appearance, particle/aggregate profile (DLS/SEC), excipient content, filter integrity (when applicable).

For lysins / depolymerases

• Identity: Peptide mapping (LC-MS/MS), intact mass.
• Potency: Kinetic lysis/turbidity assays (defined Target Kill Units), MIC/time-kill; depolymerase capsule-clearing potency (e.g., viscosity reduction, EPS sugar release).
• Impurities & safety: Endotoxin (USP <85>/<86>), HCP, residual DNA, aggregates (SEC-MALS), host strain markers (qPCR).
• Stability-indicating: Oxidation/deamidation (LC-MS), subvisible particles, activity retention vs. temp/light/freeze-thaw.

Stability programs aligned to IND/CTA

We build stability to ICH Q1A(R2): long-term and accelerated conditions; protocol includes potency maintenance, purity, aggregation, endotoxin drift, and container-closure performance. For phage DP, we often run 2–8 °C long-term plus defined stress (e.g., 25 °C/60%RH, freeze–thaw), and—if lyophilized—room-temperature brackets; enzymes follow protein-centric designs.

Data packages include protocol, timepoints, methods, acceptance criteria, and trend analysis—ready for Module 3.2.P.8.

IND (US) / CTA (EU) support

• Regulatory position. Therapeutic phage products are CBER/OVRR-regulated biological drugs; use under IND or expanded access SPIND is the current US pathway pending licensure. We prepare the CMC, environmental assessments when needed, and risk-based control strategies that tie into clinical designs.
• EMA alignment. We incorporate the EMA concept paper direction on phage-specific terminology, bank definition (phage & host cell banks), impurity control, and potency assay qualification.
• Clinical analytics. We advise on shedding/serology when relevant (virus/bacteria-based product precedents) and on sampling plans consistent with prior viral/microbial guidance.

Digital QMS, data integrity, and PPQ/CPV

All Mika facilities run an eBatch / ALCOA+ backbone. We establish scale-down models, define CPP/CQA ranges with DoE, then execute three-batch PPQ and convert to CPV dashboards for lifecycle control. EU Annex 1 environmental monitoring is trended continuously; raw data are inspection-ready.

Typical scopes & timelines (illustrative)

1. Feasibility (6–10 weeks, non-GMP): Host-range/EOP mapping; early DSP trials; enzyme potency screening; genome suitability.
2. Process development (8–16 weeks): USP/DSP formulation screens; assay validation; draft control strategy.
3. GMP DS/DP (8–12 weeks): GMP batches; release testing; stability initiation; aseptic fill/finish (vials/PFS).
4. Regulatory package: IND/CTA CMC module drafting; comparability for cocktails or sequence updates.

Interlinking services: See our Analytical & QC for Microbial Biologics page for full method menus and COA exemplars, and our Formulation & Aseptic Fill-Finish (Grade A Isolators) page for container/closure, CCIT, and lyo design options.

Why microbial-first matters here

Phages and their enzymes are, by nature, microbial products. Their upstream biology, downstream separations, and impurity risks (LPS, host DNA/protein) are different from mammalian biologics. A microbial-native CDMO will shorten the argument between bench and batch—both technically and with regulators—because the levers (host engineering, infection kinetics, nuclease windows, AEX/TFF parameters) are the ones we already hold.

You’ll notice: fewer meetings to explain fundamentals, more meetings to make decisions.

Representative release panel (phase-appropriate)

Phage DP (example)

• Identity (WGS), PFU/mL (validated), EOP on panel, sterility (USP <71>), endotoxin (USP <85>/<86>), residual DNA, residual protein, pH/osmolality, appearance, DLS/SEC profile, bioburden, filter integrity (if filtered), excipient assay.

Lysin/Depolymerase DP (example)

• Identity (LC-MS/MS), potency (turbidity/time-kill), purity (SDS-PAGE/UHPLC), endotoxin (USP <85>/<86>), HCP/HCD, residual DNA, aggregates (SEC-MALS), subvisible particles, sterility (USP <71>), stability timepoint.

Risk controls you can point to in an audit

• No lysogeny/toxins/ARGs by WGS; cocktail governance with change control.
• Endotoxin designed-out via host/media choice + AEX/TFF + acceptance criteria aligned to FDA Q&A and USP advances.
• Aseptic rigor to EU Annex 1 (2022) with Grade A isolators, Grade B background, environmental/personnel monitoring, and media fill performance trending.
• Stability to ICH Q1A(R2) with potency-preserving conditions.

Case-style outcomes

• Carbapenem-resistant Klebsiella (K64): Depolymerase + phage cocktail; EOP coverage 92% across clinical panel; depolymerase increased serum killing; DP stability 6 months at 5 °C, 1 freeze-thaw tolerated.
• MRSA bacteremia adjunct: Lysin program repositioned post-Phase 3 field lessons (trial design pitfalls, endpoints, SOC differences), with improved PK/PD sampling and potency narrative.

Start here

Send us your target organism(s), any available isolates, and your intended route/dose. We’ll propose a phase-appropriate plan with host-range milestones, DS/DSP outlines, and release panels. If you’re also exploring Bacteriocins & Lantibiotics Manufacturing (e.g., nisin, colicins) or Engineered Probiotics (LBP) GMP, we can cross-leverage analytics and suites to compress total program time.

FAQ (selected)

Q1. Are phage therapies FDA-approved? What’s the path?
A. As of today, no bacteriophage therapeutic is FDA-licensed; clinical use proceeds under IND (or Expanded Access/SPIND) with CBER/OVRR oversight. We build CMC that satisfies “safe, pure, potent, and consistently manufactured,” plan comparability for cocktail evolution, and prepare the IND/CTA package with environmental and shedding considerations where relevant.

Q2. How do you test and control endotoxin for phage and enzymes?
A. We design out LPS via host/media, apply nuclease + TFF + anion-exchange processes, and set acceptance criteria per FDA’s Pyrogen and Endotoxins Testing Q&A. For testing, we use USP <85> methods and, where validated and appropriate, USP <86> recombinant reagents (rFC/rCR) to avoid animal-derived reagents.

Q3. What’s your host-range strategy and how do you avoid overcalling it?
A. Spot tests are fast but overestimate; we quantify with EOP on panels of clinical isolates and confirm in liquid-culture kill curves. Cocktail composition is optimized for non-overlapping receptor usage and resistance-escape suppression.

Q4. Release testing—what exactly goes on the COA?
A. Phage: identity (genome), titer (PFU/mL), EOP on defined panel (if relevant), sterility (USP <71>), endotoxin (USP <85>/<86>), residual DNA/protein, appearance, pH/osmolality, particle/aggregate profile.
Enzymes: identity (LC-MS), potency (activity units), purity, endotoxin, HCP/DNA, aggregates, sterility, subvisible particles.

Q5. Stability—how durable are phages and lysins?
A. We run conditions per ICH Q1A(R2). Many phages maintain potency for months at 2–8 °C with protective excipients; lyophilization enables transport-friendly shelf life. Lysins typically favor moderate pH buffers with antioxidant/tonicity support and aggregation control; we build a stability-indicating method suite around your route and container.

Q6. Can you fill syringes or just vials? What about isolators vs. RABS?
A. We fill vials, PFS, and cartridges in Grade A isolators (Grade B background), with CCIT, media fills, and EM to EU Annex 1 (2022) expectations. For phages too large for 0.22 µm filtration, we execute fully aseptic closed-system processing and validate bioburden controls.

Q7. Do lysins work on Gram-negative bacteria?
A. Native lysins act best on Gram-positives; for Gram-negatives we apply permeabilizers or engineered constructs (e.g., “innolysins”) to cross the outer membrane—validated with activity and safety assays before scale-up.

Q8. What analytical platform breadth can you provide?
A. From PFU/EOP and genome analytics to SEC-MALS, DLS, LC-MS, BET (USP <85>/<86>), USP <71> sterility, and EM/NTA where particle analytics are needed. (For a complete list and SLAs, see our Analytical & QC for Microbial Biologics page.)

Q9. How do you handle cocktail changes over time?
A. We pre-define comparability protocols (assays, acceptance ranges, matrix of coverage) so you can update one or more phages without resetting your entire CMC narrative—critical for long-tail indications and resistance drift. EMA’s manufacturing concept paper supports clear bank/control strategy thinking, which we follow.

Q10. Can you run PPQ and set up CPV for phage and enzyme platforms?
A. Yes. We establish a qualified scale-down model, define CPPs/CQAs, run three-batch PPQ, and activate CPV dashboards with ongoing EM trending per Annex 1. (See our Process Characterization, PPQ & CPV for Microbial Platforms page for detail.)

Email our team at info@mikabiologics.com